t forsythia atcc 43037 genome  (ATCC)


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    Name:
    Tannerella forsythia FDC 338 CCUG 21028A CCUG 33064 CCUG 33226 CIP 105219 JCM 10827
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    Catalog Number:
    43037
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    Structured Review

    ATCC t forsythia atcc 43037 genome
    The percentage sequence coverage of modern and ancient known virulence factor genes; the T. forsythia 92A2 sequence was used as a reference, apart from leg when <t>ATCC</t> 43037 was used as a reference; and KLIKK protease genes when our in-home determined KLIKK protease locus was used as a reference [ 10 ]

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    t forsythia atcc 43037 genome - by Bioz Stars, 2020-09
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    Images

    1) Product Images from "Analysis of oral microbiome from fossil human remains revealed the significant differences in virulence factors of modern and ancient Tannerella forsythia"

    Article Title: Analysis of oral microbiome from fossil human remains revealed the significant differences in virulence factors of modern and ancient Tannerella forsythia

    Journal: BMC Genomics

    doi: 10.1186/s12864-020-06810-9

    The percentage sequence coverage of modern and ancient known virulence factor genes; the T. forsythia 92A2 sequence was used as a reference, apart from leg when ATCC 43037 was used as a reference; and KLIKK protease genes when our in-home determined KLIKK protease locus was used as a reference [ 10 ]
    Figure Legend Snippet: The percentage sequence coverage of modern and ancient known virulence factor genes; the T. forsythia 92A2 sequence was used as a reference, apart from leg when ATCC 43037 was used as a reference; and KLIKK protease genes when our in-home determined KLIKK protease locus was used as a reference [ 10 ]

    Techniques Used: Sequencing

    DNA sequence comparison of the  T. forsythia  reference genome to the ancient  T. forsythia  genomes and to publicly available modern  T. forsythia  genomes. The two outermost rings depict the forward and reverse coding strands of the reference genome. The next 13 rings moving towards the inner part of the figure display regions of sequence similarity detected by BLAST comparison between the DNA of the reference genome and the DNA of the 13 compared  T. forsythia  genomes. The following genome order reflects the order of the circles starting from the outer part of the figure and moving towards the inner part: PCA0332, PCA0198, UB20, PCA0088, KS16, UB4, UB22, NSLJ, G12, 3313, NSLK, ATCC 43037, and 9610. Genes associated with  T. forsythia  virulence are labeled in the plot
    Figure Legend Snippet: DNA sequence comparison of the T. forsythia reference genome to the ancient T. forsythia genomes and to publicly available modern T. forsythia genomes. The two outermost rings depict the forward and reverse coding strands of the reference genome. The next 13 rings moving towards the inner part of the figure display regions of sequence similarity detected by BLAST comparison between the DNA of the reference genome and the DNA of the 13 compared T. forsythia genomes. The following genome order reflects the order of the circles starting from the outer part of the figure and moving towards the inner part: PCA0332, PCA0198, UB20, PCA0088, KS16, UB4, UB22, NSLJ, G12, 3313, NSLK, ATCC 43037, and 9610. Genes associated with T. forsythia virulence are labeled in the plot

    Techniques Used: Sequencing, Labeling

    2) Product Images from "Comparative genome characterization of the periodontal pathogen Tannerella forsythia"

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia

    Journal: BMC Genomics

    doi: 10.1186/s12864-020-6535-y

    Analysis of codon usage for ATCC 43037 (left panel) and BU063 (right panel). The continuous curves indicate the NC values to be expected for a given GC3s content in the absence of other factors shaping codon usage. Every dot represents a protein coding gene, dots not positioned near the curve therefore represent genes that display a considerable codon usage bias. GC3s: G + C content at synonymous positions, NC: effective number of codons used within the sequence of a gene
    Figure Legend Snippet: Analysis of codon usage for ATCC 43037 (left panel) and BU063 (right panel). The continuous curves indicate the NC values to be expected for a given GC3s content in the absence of other factors shaping codon usage. Every dot represents a protein coding gene, dots not positioned near the curve therefore represent genes that display a considerable codon usage bias. GC3s: G + C content at synonymous positions, NC: effective number of codons used within the sequence of a gene

    Techniques Used: Sequencing

    Multiple whole genome alignment of eight T. forsythia strains. Each coloured block represents a genomic region that aligned to a region in at least one other genome, plotted in the same colour, to which it was predicted to be homologous based on sequence similarity. Blocks above the centre line indicate forward orientation; blocks below the line indicate reverse orientation relative to strain 92A2. A histogram within each block shows the average similarity of a region to its counterparts in the other genomes. Red vertical lines indicate contig boundaries. Strain ATCC 43037 displayed two translocations compared to strain 92A2 with lengths of approximately 500 kbp (blue and yellow blocks at the right end of 92A2 and in the centre of ATCC) and 30 kbp (pink block at approx. 1.25 Mbp in 92A2 and at approx. 2.7 Mbp in ATCC), respectively. Previously described large-scale inversions in strain KS16 could be confirmed (reverted blocks in the left half of the alignment)
    Figure Legend Snippet: Multiple whole genome alignment of eight T. forsythia strains. Each coloured block represents a genomic region that aligned to a region in at least one other genome, plotted in the same colour, to which it was predicted to be homologous based on sequence similarity. Blocks above the centre line indicate forward orientation; blocks below the line indicate reverse orientation relative to strain 92A2. A histogram within each block shows the average similarity of a region to its counterparts in the other genomes. Red vertical lines indicate contig boundaries. Strain ATCC 43037 displayed two translocations compared to strain 92A2 with lengths of approximately 500 kbp (blue and yellow blocks at the right end of 92A2 and in the centre of ATCC) and 30 kbp (pink block at approx. 1.25 Mbp in 92A2 and at approx. 2.7 Mbp in ATCC), respectively. Previously described large-scale inversions in strain KS16 could be confirmed (reverted blocks in the left half of the alignment)

    Techniques Used: Blocking Assay, Sequencing

    Whole genome alignment between the six frame amino acid translations of both Tannerella sp. BU063 and the scaffolded and ordered ATCC 43037 assembly. Whereas the amino acid alignment reflects similarity with respect to gene content, the order of genes is not preserved
    Figure Legend Snippet: Whole genome alignment between the six frame amino acid translations of both Tannerella sp. BU063 and the scaffolded and ordered ATCC 43037 assembly. Whereas the amino acid alignment reflects similarity with respect to gene content, the order of genes is not preserved

    Techniques Used:

    3) Product Images from "A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications"

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.02008

    (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representations ( Varki et al., 2015 ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.
    Figure Legend Snippet: (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representations ( Varki et al., 2015 ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.

    Techniques Used: Staining, Labeling, Mass Spectrometry, Molecular Weight, Marker, Western Blot, Migration

    Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.
    Figure Legend Snippet: Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.

    Techniques Used: Sequencing

    ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representation ( Varki et al., 2015 ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.
    Figure Legend Snippet: ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representation ( Varki et al., 2015 ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.

    Techniques Used: Mass Spectrometry, Knock-Out

    (A) Scheme of the T. forsythia ATCC 43037 S-layer O -glycan structure. Monosaccharide symbols are shown according to the Symbol Nomenclature for Glycans (SNFG) ( Varki et al., 2015 ). Please note that the position of the branching Fuc remained unclear ( Posch et al., 2011 ) until it was determined in the course of this study to be on the reducing-end Gal. (B) Scheme of the 27-kb protein O -glycosylation gene cluster of T. forsythia ATCC 43037. Wzx (black), flippase; pseBCFHGI (green), CMP-Pse biosynthesis genes; gtfSMILE (blue), Gtf genes; mtfJOY (yellow), Mtf genes; asnB (putative asparagine synthetase B), wecC (UDP- N -acetyl- D -mannosamine dehydrogenase) and wecB (UDP- N- acetylglucosamine 2-epimerase) (purple); hypothetical proteins, HP (gray). Genes are not drawn to scale.
    Figure Legend Snippet: (A) Scheme of the T. forsythia ATCC 43037 S-layer O -glycan structure. Monosaccharide symbols are shown according to the Symbol Nomenclature for Glycans (SNFG) ( Varki et al., 2015 ). Please note that the position of the branching Fuc remained unclear ( Posch et al., 2011 ) until it was determined in the course of this study to be on the reducing-end Gal. (B) Scheme of the 27-kb protein O -glycosylation gene cluster of T. forsythia ATCC 43037. Wzx (black), flippase; pseBCFHGI (green), CMP-Pse biosynthesis genes; gtfSMILE (blue), Gtf genes; mtfJOY (yellow), Mtf genes; asnB (putative asparagine synthetase B), wecC (UDP- N -acetyl- D -mannosamine dehydrogenase) and wecB (UDP- N- acetylglucosamine 2-epimerase) (purple); hypothetical proteins, HP (gray). Genes are not drawn to scale.

    Techniques Used:

    Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.
    Figure Legend Snippet: Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.

    Techniques Used: Methylation

    4) Product Images from "Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications"

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    Journal: Glycobiology

    doi: 10.1093/glycob/cww129

    Knockout of pseC and legC decreases biofilm formation of T. forsythia ATCC 43037 and T. forsythia UB4 cells, respectively. ( A ) Biofilm formation in T. forsythia ATCC 43037 wild-type compared to ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp . ( B ) Biofilm formation in T. forsythia UB4 wild-type compared to UB4 Δ legC and the complemented strain T. forsythia UB4 Δ legC comp . Data represent mean values ± SD of at least four independent experiments with four replicates each and were analyzed by the unpaired Student’s t-test. Asterisks indicate significant differences (** P
    Figure Legend Snippet: Knockout of pseC and legC decreases biofilm formation of T. forsythia ATCC 43037 and T. forsythia UB4 cells, respectively. ( A ) Biofilm formation in T. forsythia ATCC 43037 wild-type compared to ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp . ( B ) Biofilm formation in T. forsythia UB4 wild-type compared to UB4 Δ legC and the complemented strain T. forsythia UB4 Δ legC comp . Data represent mean values ± SD of at least four independent experiments with four replicates each and were analyzed by the unpaired Student’s t-test. Asterisks indicate significant differences (** P

    Techniques Used: Knock-Out

    ( A ) Structures of Sia and Sia-like sugars (NulOs). Sialic acid (Neu5Ac), legionaminic acid (Leg5,7Ac 2 ) and pseudaminic acids (Pse5,7Ac 2 and Pse5Am7Gra) are shown. To note, Pse5Am7Gra is found as the terminal sugar of the S-layer glycan in T. forsythia ATCC 43037. For reference, the nine carbon atoms of Sia are numbered, and the structure of the NHAc group is shown in the boxed inset. ( B ) Schematic drawing of the structure of the S-layer O -glycan in T. forsythia ATCC 43037 (amended from Posch et al. (2011) ). This figure is available in black and white in print and in color at Glycobiology online.
    Figure Legend Snippet: ( A ) Structures of Sia and Sia-like sugars (NulOs). Sialic acid (Neu5Ac), legionaminic acid (Leg5,7Ac 2 ) and pseudaminic acids (Pse5,7Ac 2 and Pse5Am7Gra) are shown. To note, Pse5Am7Gra is found as the terminal sugar of the S-layer glycan in T. forsythia ATCC 43037. For reference, the nine carbon atoms of Sia are numbered, and the structure of the NHAc group is shown in the boxed inset. ( B ) Schematic drawing of the structure of the S-layer O -glycan in T. forsythia ATCC 43037 (amended from Posch et al. (2011) ). This figure is available in black and white in print and in color at Glycobiology online.

    Techniques Used:

    Experimentally confirmed enzymatic steps (in green) in T. forsythia strains ATCC 43037 (left) and FDC 92A2/UB4 (right) corresponding to the CMP-Pse and CMP-Leg biosynthetic pathways as elucidated in H. pylori and C. jejuni , respectively ( Schoenhofen et al. 2006b , Schoenhofen et al. 2009 ). In T. forsythia , pathways will deviate at some point to produce the unique NulO derivatives found in our strains, such as Pse5Am7Gra. These deviations would be anticipated to occur either within the NulO biosynthetic pathway or post CMP-NulO biosynthesis. Red, blue and orange highlight enzymatic steps that introduce the stereochemical differences between the two pathways and also indicate the positions altered for both hexose intermediates and final NulO. The assignment of roman numerals to each compound is consistent with label designations found throughout the text. Subscripts “P” and “L” indicate intermediates from the CMP-Pse and CMP-Leg biosynthesis pathway, respectively. For simplicity, all sugars except for the NulOs are shown in 4 C 1 form. This figure is available in black and white in print and in color at Glycobiology online.
    Figure Legend Snippet: Experimentally confirmed enzymatic steps (in green) in T. forsythia strains ATCC 43037 (left) and FDC 92A2/UB4 (right) corresponding to the CMP-Pse and CMP-Leg biosynthetic pathways as elucidated in H. pylori and C. jejuni , respectively ( Schoenhofen et al. 2006b , Schoenhofen et al. 2009 ). In T. forsythia , pathways will deviate at some point to produce the unique NulO derivatives found in our strains, such as Pse5Am7Gra. These deviations would be anticipated to occur either within the NulO biosynthetic pathway or post CMP-NulO biosynthesis. Red, blue and orange highlight enzymatic steps that introduce the stereochemical differences between the two pathways and also indicate the positions altered for both hexose intermediates and final NulO. The assignment of roman numerals to each compound is consistent with label designations found throughout the text. Subscripts “P” and “L” indicate intermediates from the CMP-Pse and CMP-Leg biosynthesis pathway, respectively. For simplicity, all sugars except for the NulOs are shown in 4 C 1 form. This figure is available in black and white in print and in color at Glycobiology online.

    Techniques Used: Introduce

    Phylogenetic clusters of selected microbial NulO synthase homologs. Based on the prediction of NulO sugar type by Lewis et al. (2009) , a distance-based neighbor joining tree calculated from the amino acid sequences of NulO synthase enzymes places Tanf_01240 of T. forsythia type strain ATCC 43037 in a phylogenetic clade of pseudaminic acid synthases, while BFO_1066 of strain FDC 92A2/UB4 is clustered with Leg synthases (bootstrap values shown at relevant nodes). Green denotes Leg synthases, red Neu synthases and blue Pse synthases. This figure is available in black and white in print and in color at Glycobiology online.
    Figure Legend Snippet: Phylogenetic clusters of selected microbial NulO synthase homologs. Based on the prediction of NulO sugar type by Lewis et al. (2009) , a distance-based neighbor joining tree calculated from the amino acid sequences of NulO synthase enzymes places Tanf_01240 of T. forsythia type strain ATCC 43037 in a phylogenetic clade of pseudaminic acid synthases, while BFO_1066 of strain FDC 92A2/UB4 is clustered with Leg synthases (bootstrap values shown at relevant nodes). Green denotes Leg synthases, red Neu synthases and blue Pse synthases. This figure is available in black and white in print and in color at Glycobiology online.

    Techniques Used:

    SDS-PAGE gels (12%) of purified recombinant enzymes of the ( A ) CMP-Pse biosynthesis pathway from T. forsythia ATCC 43037 and ( B ) CMP-Leg biosynthesis pathway from T. forsythia FDC 92A2/UB4.
    Figure Legend Snippet: SDS-PAGE gels (12%) of purified recombinant enzymes of the ( A ) CMP-Pse biosynthesis pathway from T. forsythia ATCC 43037 and ( B ) CMP-Leg biosynthesis pathway from T. forsythia FDC 92A2/UB4.

    Techniques Used: SDS Page, Purification, Recombinant

    SDS-PAGE analysis of T. forsythia parent, NulO-deficient, and complemented strains. ( A ) CBB stained SDS-PAGE (7.5% gel) of whole cell extracts from T. forsythia ATCC 43037, ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp , and ( B ) T. forsythia UB4 wild-type, UB4 Δ legC and the complemented strain UB4 Δ legC comp . For both T. forsythia ATCC 43037 Δ pseC and UB4 Δ legC , a downshift of the S-layer protein bands (TfsA and TfsB) could be observed, which was reverted in the complemented strains.
    Figure Legend Snippet: SDS-PAGE analysis of T. forsythia parent, NulO-deficient, and complemented strains. ( A ) CBB stained SDS-PAGE (7.5% gel) of whole cell extracts from T. forsythia ATCC 43037, ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp , and ( B ) T. forsythia UB4 wild-type, UB4 Δ legC and the complemented strain UB4 Δ legC comp . For both T. forsythia ATCC 43037 Δ pseC and UB4 Δ legC , a downshift of the S-layer protein bands (TfsA and TfsB) could be observed, which was reverted in the complemented strains.

    Techniques Used: SDS Page, Staining

    5) Product Images from "A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications"

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.02008

    (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representations ( Varki et al., 2015 ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.
    Figure Legend Snippet: (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representations ( Varki et al., 2015 ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.

    Techniques Used: Staining, Labeling, Mass Spectrometry, Molecular Weight, Marker, Western Blot, Migration

    Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.
    Figure Legend Snippet: Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.

    Techniques Used: Sequencing

    ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representation ( Varki et al., 2015 ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.
    Figure Legend Snippet: ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z values) are shown in SNFG representation ( Varki et al., 2015 ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.

    Techniques Used: Mass Spectrometry, Knock-Out

    (A) Scheme of the T. forsythia ATCC 43037 S-layer O -glycan structure. Monosaccharide symbols are shown according to the Symbol Nomenclature for Glycans (SNFG) ( Varki et al., 2015 ). Please note that the position of the branching Fuc remained unclear ( Posch et al., 2011 ) until it was determined in the course of this study to be on the reducing-end Gal. (B) Scheme of the 27-kb protein O -glycosylation gene cluster of T. forsythia ATCC 43037. Wzx (black), flippase; pseBCFHGI (green), CMP-Pse biosynthesis genes; gtfSMILE (blue), Gtf genes; mtfJOY (yellow), Mtf genes; asnB (putative asparagine synthetase B), wecC (UDP- N -acetyl- D -mannosamine dehydrogenase) and wecB (UDP- N- acetylglucosamine 2-epimerase) (purple); hypothetical proteins, HP (gray). Genes are not drawn to scale.
    Figure Legend Snippet: (A) Scheme of the T. forsythia ATCC 43037 S-layer O -glycan structure. Monosaccharide symbols are shown according to the Symbol Nomenclature for Glycans (SNFG) ( Varki et al., 2015 ). Please note that the position of the branching Fuc remained unclear ( Posch et al., 2011 ) until it was determined in the course of this study to be on the reducing-end Gal. (B) Scheme of the 27-kb protein O -glycosylation gene cluster of T. forsythia ATCC 43037. Wzx (black), flippase; pseBCFHGI (green), CMP-Pse biosynthesis genes; gtfSMILE (blue), Gtf genes; mtfJOY (yellow), Mtf genes; asnB (putative asparagine synthetase B), wecC (UDP- N -acetyl- D -mannosamine dehydrogenase) and wecB (UDP- N- acetylglucosamine 2-epimerase) (purple); hypothetical proteins, HP (gray). Genes are not drawn to scale.

    Techniques Used:

    Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.
    Figure Legend Snippet: Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.

    Techniques Used: Methylation

    6) Product Images from "A pseudaminic acid or a legionaminic acid derivative transferase is strain-specifically implicated in the general protein O-glycosylation system of the periodontal pathogen Tannerella forsythia"

    Article Title: A pseudaminic acid or a legionaminic acid derivative transferase is strain-specifically implicated in the general protein O-glycosylation system of the periodontal pathogen Tannerella forsythia

    Journal: Glycobiology

    doi: 10.1093/glycob/cwx019

    Deconvoluted ESI-IT-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia parent and mutant strains. ( A ) Comparison of the spectra from T. forsythia ATCC 43037 wild-type, Δ Tanf_01245 mutant, reconstituted mutant Δ Tanf_01245 + and cross-complemented mutant Δ Tanf_01245 + TFUB4_00887 . ( B ) Comparison of the spectra from T. forsythia UB4 wild-type, Δ TFUB4_00887 mutant, reconstituted mutant Δ TFUB4_00887 + and cross-complemented mutant Δ TFUB4_00887 + Tanf_01245 . Another glycan species with additional +16 Da at the position of the digitoxose was observed, indicative of the presence of a deoxyhexose instead of a dideoxyhexose in some forms of the glycan. The glycan structures of the highest mass peaks are shown as symbolic representations. Mass peaks from the subsequent fragmentation pattern were assigned according to the loss of carbohydrate units and modifications. Relative peak intensities of occurring peaks are given on the y axis. This figure is available in black and white in print and in color at Glycobiology online.
    Figure Legend Snippet: Deconvoluted ESI-IT-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia parent and mutant strains. ( A ) Comparison of the spectra from T. forsythia ATCC 43037 wild-type, Δ Tanf_01245 mutant, reconstituted mutant Δ Tanf_01245 + and cross-complemented mutant Δ Tanf_01245 + TFUB4_00887 . ( B ) Comparison of the spectra from T. forsythia UB4 wild-type, Δ TFUB4_00887 mutant, reconstituted mutant Δ TFUB4_00887 + and cross-complemented mutant Δ TFUB4_00887 + Tanf_01245 . Another glycan species with additional +16 Da at the position of the digitoxose was observed, indicative of the presence of a deoxyhexose instead of a dideoxyhexose in some forms of the glycan. The glycan structures of the highest mass peaks are shown as symbolic representations. Mass peaks from the subsequent fragmentation pattern were assigned according to the loss of carbohydrate units and modifications. Relative peak intensities of occurring peaks are given on the y axis. This figure is available in black and white in print and in color at Glycobiology online.

    Techniques Used: Mass Spectrometry, Mutagenesis

    ESI-IT-MS analysis of cellular nucleotide sugar pools from T. forsythia strains. ( A ) CMP-activated Pse5Am7Gra ( m / z 683.3) was detected in the T. forsythia ATCC 43037 wild-type and in the Δ Tanf_01245 mutant, whereas this mass was absent in a Pse biosynthesis deficient strain (Δ pseC ) which served as a negative control. ( B ) In T. forsythia UB4 wild-type and in the Δ TFUB4_00887 mutant, a m / z 654.3 peak was identified, which was attributed to a CMP-activated Leg derivative (CMP-Leg*). This mass is consistent with having Ac and Gc modifications on Leg, based on calculation. Notably, this peak was absent in the Legbiosynthesis deficient strain (Δ legC ) which served as a negative control. Relative peak intensities are given on the y axis.
    Figure Legend Snippet: ESI-IT-MS analysis of cellular nucleotide sugar pools from T. forsythia strains. ( A ) CMP-activated Pse5Am7Gra ( m / z 683.3) was detected in the T. forsythia ATCC 43037 wild-type and in the Δ Tanf_01245 mutant, whereas this mass was absent in a Pse biosynthesis deficient strain (Δ pseC ) which served as a negative control. ( B ) In T. forsythia UB4 wild-type and in the Δ TFUB4_00887 mutant, a m / z 654.3 peak was identified, which was attributed to a CMP-activated Leg derivative (CMP-Leg*). This mass is consistent with having Ac and Gc modifications on Leg, based on calculation. Notably, this peak was absent in the Legbiosynthesis deficient strain (Δ legC ) which served as a negative control. Relative peak intensities are given on the y axis.

    Techniques Used: Mass Spectrometry, Mutagenesis, Negative Control

    SDS-PAGE and Western immunoblot analyses of T. forsythia ATCC 43037 and T. forsythia UB4 wild-type and mutants. ( A ) CBB staining of crude cell extracts from T. forsythia ATCC 43037 wild-type, Δ Tanf_01245 mutant, reconstituted mutant Δ Tanf_01245 + and cross-complemented mutant Δ Tanf_01245 + TFUB4_00887 after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the down-shift resulting from the loss of the Pse5Am7Gra residue can be observed in the deletion mutant and in the cross-complemented mutant, while in the reconstituted strain the bands are up-shifted again to wild-type level. The same migration profiles could be observed for T. forsythia UB4 wild-type, Δ TFUB4_00887 mutant, reconstituted mutant Δ TFUB4_00887 + and cross-complemented mutant Δ TFUB4_00887 + Tanf_01245 . PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. The S-layer glycoprotein bands were further processed for MS analyses. Western immunoblots probed with anti-TfsA antiserum ( B ) and anti-TfsB antiserum ( C ) confirmed the identity of the S-layer glycoproteins in all analyzed T. forsythia species. PageRuler Prestained Protein Ladder (Thermo Fisher Scientific) was used as a molecular weight marker.
    Figure Legend Snippet: SDS-PAGE and Western immunoblot analyses of T. forsythia ATCC 43037 and T. forsythia UB4 wild-type and mutants. ( A ) CBB staining of crude cell extracts from T. forsythia ATCC 43037 wild-type, Δ Tanf_01245 mutant, reconstituted mutant Δ Tanf_01245 + and cross-complemented mutant Δ Tanf_01245 + TFUB4_00887 after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the down-shift resulting from the loss of the Pse5Am7Gra residue can be observed in the deletion mutant and in the cross-complemented mutant, while in the reconstituted strain the bands are up-shifted again to wild-type level. The same migration profiles could be observed for T. forsythia UB4 wild-type, Δ TFUB4_00887 mutant, reconstituted mutant Δ TFUB4_00887 + and cross-complemented mutant Δ TFUB4_00887 + Tanf_01245 . PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. The S-layer glycoprotein bands were further processed for MS analyses. Western immunoblots probed with anti-TfsA antiserum ( B ) and anti-TfsB antiserum ( C ) confirmed the identity of the S-layer glycoproteins in all analyzed T. forsythia species. PageRuler Prestained Protein Ladder (Thermo Fisher Scientific) was used as a molecular weight marker.

    Techniques Used: SDS Page, Western Blot, Staining, Mutagenesis, Labeling, Migration, Molecular Weight, Marker, Mass Spectrometry

    7) Product Images from "A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications"

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.02008

    (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.
    Figure Legend Snippet: (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.

    Techniques Used: Staining, Labeling, Mass Spectrometry, Molecular Weight, Marker, Western Blot, Migration

    Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.
    Figure Legend Snippet: Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.

    Techniques Used: Sequencing

    ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.
    Figure Legend Snippet: ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.

    Techniques Used: Mass Spectrometry, Knock-Out

    Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.
    Figure Legend Snippet: Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.

    Techniques Used: Methylation

    8) Product Images from "A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications"

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.02008

    (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.
    Figure Legend Snippet: (A) Coomassie Brilliant Blue staining of crude cell extracts from T. forsythia ATCC 43037 wild-type and glycosyltransferase-deficient mutants after separation on a 7.5% SDS-PA gel. The S-layer glycoproteins (labeled TfsA and TfsB) are indicated and the downshifts resulting from glycan truncation can be seen in the mutants. S-layer glycoprotein bands were further processed for MS analyses. PageRuler Plus Prestained Protein Ladder (Thermo Fisher Scientific) was used as a protein molecular weight marker. (B) Western-blots probed with α-TfsA and α-TfsB antiserum for confirmation of the identity of S-layer glycoproteins. Glycoproteins from all glycosyltransferase-deficient mutants (Δ gtfSMILE ) experienced a downshift resulting from glycan truncation, whereas the reconstituted strains (denoted with +) regained wild-type migration, indicating the presence of the complete mature glycan, proving successful recombination. (C, i–vi) ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia wild-type and mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). O -Glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications.

    Techniques Used: Staining, Labeling, Mass Spectrometry, Molecular Weight, Marker, Western Blot, Migration

    Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.
    Figure Legend Snippet: Alignment of protein O- glycosylation gene clusters from different T. forsythia strains showing comparable sizes and gene organizations (drawn to scale). Genes showing sequence identity > 50% and sequence coverage > 50% between strains appear in the same color. The major difference in the analyzed strains are for genes synthesizing either CMP-Pse ( pseBCFHGI ; light green colors; strains ATCC 43037 and UB20) or CMP-Leg ( legBCHIGF, ptmE ; dark green colors; strains FDC 92A2, UB4, KS16, UB22). Genes encoding Gtfs ( gtfSMILE ; blue color), Mtfs ( mtfJOY ; yellow color) and carbohydrate modifying enzymes ( asnB, wecC, wecB ; gray color) show high sequence homology between analyzed strains. Genomes of all strains synthesizing CMP-Leg encode an additional putative Mtf gene ( mtfX ), which does not share sequence homology to other Mtfs located within the cluster. In strain UB22, mtfJ is not predicted and for strain 3313 only five out of seven genes needed for the synthesis of CMP-Leg are predicted confidently. Due to low homology, isolate Tannerella sp. HOT-286 (phylotype BU063) could not be aligned with the other T. forsythia strains; for that isolate, the genomic area between a wzx -like gene and the gtfE gene is shown for comparison. P, Pse transferase; L, Leg transferase; HP, hypothetical protein; the star symbol ( ∗ ) indicates a transposable element; genes written in bold letters were investigated in detail in course of this study.

    Techniques Used: Sequencing

    ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.
    Figure Legend Snippet: ESI-MS sum spectra of β-eliminated TfsB O -glycans from T. forsythia ATCC 43037 methyltransferase knock-out mutants. The glycan structures of the signals corresponding to the largest mass (bold m / z ). Other O -glycan signals detected for the respective mutants were assigned based on the m / z mass differences corresponding to the loss of individual sugar units and/or modifications. The lack of methyl modifications is indicated by a red circle in the symbolic O -glycan structure representation.

    Techniques Used: Mass Spectrometry, Knock-Out

    Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.
    Figure Legend Snippet: Model for the biosynthesis of the species-specific portion of the T. forsythia ATCC 43037 O -glycan. Upon synthesis of the pentasaccharide core on an undP lipid carrier, the first carbohydrate residue of the species-specific glycan is a Fuc residue conferred by GtfE. The glycan is elongated with a Gal residue which is transferred by GtfL and methylated by MtfY. The assembly of the three sugar branch, consisting of a ManNAcA residue (transferred by GtfI), a ManNAcCONH 2 residue (GtfM), which is methylated by either MtfJ or MtfO, and a Pse5Am7Gra residue (transferred via GtfS), completes the synthesis of the decasaccharide.

    Techniques Used: Methylation

    Related Articles

    Sequencing:

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia
    Article Snippet: .. Improved assembly of the Tannerella forsythia type strain ATCC 43037 The genome of the T. forsythia ATCC 43037 type strain had been assembled previously [ ] based on Illumina paired-end sequencing data resulting in an assembly of 141 contigs with an N50 size of 114 kilobasepairs (kbp) (Table ). .. The largest sequence was 487 kbp comprising about 15% of the total assembly size of 3.282 Megabasepairs (Mbp).

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia
    Article Snippet: .. Mate-pairs of T. forsythia ATCC 43037 were deposited in the Sequence Read Archive under accession SRR9302598 (BioProject PRJNA548889, BioSample SAMN12058270). ..

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia
    Article Snippet: .. Previous characterizations of the T. forsythia virulence factors were mostly based on the American Type Culture Collection (ATCC) 43037 type strain employing wet-lab experimentation, whereas computational analyses of the virulence-related gene repertoire mostly used the genome sequence of strain FDC 92A2. .. Although FDC 92A2 was the first fully sequenced T. forsythia strain available [ ], the resulting genome assembly was incorrectly labelled and deposited as ATCC 43037 in the National Center for Biotechnology Information (NCBI) databases.

    Mutagenesis:

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications
    Article Snippet: .. In comparison to the T. forsythia ATCC 43037 wild-type, in which the glycosylated S-layer proteins TfsA and TfsB migrate on the SDS-PAGE gel at ∼230 kDa (calculated MW of the protein, 135 kDa) and ∼270 kDa (calculated MW of the protein, 152 kDa), respectively, each Gtf-deficient mutant experienced a downshift of these prominent T. forsythia glycoproteins ( Figure ). ..

    SDS Page:

    Article Title: A General Protein O-Glycosylation Gene Cluster Encodes the Species-Specific Glycan of the Oral Pathogen Tannerella forsythia: O-Glycan Biosynthesis and Immunological Implications
    Article Snippet: .. In comparison to the T. forsythia ATCC 43037 wild-type, in which the glycosylated S-layer proteins TfsA and TfsB migrate on the SDS-PAGE gel at ∼230 kDa (calculated MW of the protein, 135 kDa) and ∼270 kDa (calculated MW of the protein, 152 kDa), respectively, each Gtf-deficient mutant experienced a downshift of these prominent T. forsythia glycoproteins ( Figure ). ..

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    ATCC t forsythia atcc 43037 type strain
    Analysis of codon usage for <t>ATCC</t> 43037 (left panel) and BU063 (right panel). The continuous curves indicate the NC values to be expected for a given GC3s content in the absence of other factors shaping codon usage. Every dot represents a protein coding gene, dots not positioned near the curve therefore represent genes that display a considerable codon usage bias. GC3s: G + C content at synonymous positions, NC: effective number of codons used within the sequence of a gene
    T Forsythia Atcc 43037 Type Strain, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Analysis of codon usage for ATCC 43037 (left panel) and BU063 (right panel). The continuous curves indicate the NC values to be expected for a given GC3s content in the absence of other factors shaping codon usage. Every dot represents a protein coding gene, dots not positioned near the curve therefore represent genes that display a considerable codon usage bias. GC3s: G + C content at synonymous positions, NC: effective number of codons used within the sequence of a gene

    Journal: BMC Genomics

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia

    doi: 10.1186/s12864-020-6535-y

    Figure Lengend Snippet: Analysis of codon usage for ATCC 43037 (left panel) and BU063 (right panel). The continuous curves indicate the NC values to be expected for a given GC3s content in the absence of other factors shaping codon usage. Every dot represents a protein coding gene, dots not positioned near the curve therefore represent genes that display a considerable codon usage bias. GC3s: G + C content at synonymous positions, NC: effective number of codons used within the sequence of a gene

    Article Snippet: Improved assembly of the Tannerella forsythia type strain ATCC 43037 The genome of the T. forsythia ATCC 43037 type strain had been assembled previously [ ] based on Illumina paired-end sequencing data resulting in an assembly of 141 contigs with an N50 size of 114 kilobasepairs (kbp) (Table ).

    Techniques: Sequencing

    Multiple whole genome alignment of eight T. forsythia strains. Each coloured block represents a genomic region that aligned to a region in at least one other genome, plotted in the same colour, to which it was predicted to be homologous based on sequence similarity. Blocks above the centre line indicate forward orientation; blocks below the line indicate reverse orientation relative to strain 92A2. A histogram within each block shows the average similarity of a region to its counterparts in the other genomes. Red vertical lines indicate contig boundaries. Strain ATCC 43037 displayed two translocations compared to strain 92A2 with lengths of approximately 500 kbp (blue and yellow blocks at the right end of 92A2 and in the centre of ATCC) and 30 kbp (pink block at approx. 1.25 Mbp in 92A2 and at approx. 2.7 Mbp in ATCC), respectively. Previously described large-scale inversions in strain KS16 could be confirmed (reverted blocks in the left half of the alignment)

    Journal: BMC Genomics

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia

    doi: 10.1186/s12864-020-6535-y

    Figure Lengend Snippet: Multiple whole genome alignment of eight T. forsythia strains. Each coloured block represents a genomic region that aligned to a region in at least one other genome, plotted in the same colour, to which it was predicted to be homologous based on sequence similarity. Blocks above the centre line indicate forward orientation; blocks below the line indicate reverse orientation relative to strain 92A2. A histogram within each block shows the average similarity of a region to its counterparts in the other genomes. Red vertical lines indicate contig boundaries. Strain ATCC 43037 displayed two translocations compared to strain 92A2 with lengths of approximately 500 kbp (blue and yellow blocks at the right end of 92A2 and in the centre of ATCC) and 30 kbp (pink block at approx. 1.25 Mbp in 92A2 and at approx. 2.7 Mbp in ATCC), respectively. Previously described large-scale inversions in strain KS16 could be confirmed (reverted blocks in the left half of the alignment)

    Article Snippet: Improved assembly of the Tannerella forsythia type strain ATCC 43037 The genome of the T. forsythia ATCC 43037 type strain had been assembled previously [ ] based on Illumina paired-end sequencing data resulting in an assembly of 141 contigs with an N50 size of 114 kilobasepairs (kbp) (Table ).

    Techniques: Blocking Assay, Sequencing

    Whole genome alignment between the six frame amino acid translations of both Tannerella sp. BU063 and the scaffolded and ordered ATCC 43037 assembly. Whereas the amino acid alignment reflects similarity with respect to gene content, the order of genes is not preserved

    Journal: BMC Genomics

    Article Title: Comparative genome characterization of the periodontal pathogen Tannerella forsythia

    doi: 10.1186/s12864-020-6535-y

    Figure Lengend Snippet: Whole genome alignment between the six frame amino acid translations of both Tannerella sp. BU063 and the scaffolded and ordered ATCC 43037 assembly. Whereas the amino acid alignment reflects similarity with respect to gene content, the order of genes is not preserved

    Article Snippet: Improved assembly of the Tannerella forsythia type strain ATCC 43037 The genome of the T. forsythia ATCC 43037 type strain had been assembled previously [ ] based on Illumina paired-end sequencing data resulting in an assembly of 141 contigs with an N50 size of 114 kilobasepairs (kbp) (Table ).

    Techniques:

    The percentage sequence coverage of modern and ancient known virulence factor genes; the T. forsythia 92A2 sequence was used as a reference, apart from leg when ATCC 43037 was used as a reference; and KLIKK protease genes when our in-home determined KLIKK protease locus was used as a reference [ 10 ]

    Journal: BMC Genomics

    Article Title: Analysis of oral microbiome from fossil human remains revealed the significant differences in virulence factors of modern and ancient Tannerella forsythia

    doi: 10.1186/s12864-020-06810-9

    Figure Lengend Snippet: The percentage sequence coverage of modern and ancient known virulence factor genes; the T. forsythia 92A2 sequence was used as a reference, apart from leg when ATCC 43037 was used as a reference; and KLIKK protease genes when our in-home determined KLIKK protease locus was used as a reference [ 10 ]

    Article Snippet: Sequencing of the T. forsythia ATCC 43037 genome, apart from bspA (BFO_RS14480 ), revealed five more genes (BFO_RS14345 (bspB ), BFO_RS08355 , BFO_RS14330 , BFO_RS14330 , and BFO_RS14330 ) encoding putative BspA-like proteins.

    Techniques: Sequencing

    DNA sequence comparison of the  T. forsythia  reference genome to the ancient  T. forsythia  genomes and to publicly available modern  T. forsythia  genomes. The two outermost rings depict the forward and reverse coding strands of the reference genome. The next 13 rings moving towards the inner part of the figure display regions of sequence similarity detected by BLAST comparison between the DNA of the reference genome and the DNA of the 13 compared  T. forsythia  genomes. The following genome order reflects the order of the circles starting from the outer part of the figure and moving towards the inner part: PCA0332, PCA0198, UB20, PCA0088, KS16, UB4, UB22, NSLJ, G12, 3313, NSLK, ATCC 43037, and 9610. Genes associated with  T. forsythia  virulence are labeled in the plot

    Journal: BMC Genomics

    Article Title: Analysis of oral microbiome from fossil human remains revealed the significant differences in virulence factors of modern and ancient Tannerella forsythia

    doi: 10.1186/s12864-020-06810-9

    Figure Lengend Snippet: DNA sequence comparison of the T. forsythia reference genome to the ancient T. forsythia genomes and to publicly available modern T. forsythia genomes. The two outermost rings depict the forward and reverse coding strands of the reference genome. The next 13 rings moving towards the inner part of the figure display regions of sequence similarity detected by BLAST comparison between the DNA of the reference genome and the DNA of the 13 compared T. forsythia genomes. The following genome order reflects the order of the circles starting from the outer part of the figure and moving towards the inner part: PCA0332, PCA0198, UB20, PCA0088, KS16, UB4, UB22, NSLJ, G12, 3313, NSLK, ATCC 43037, and 9610. Genes associated with T. forsythia virulence are labeled in the plot

    Article Snippet: Sequencing of the T. forsythia ATCC 43037 genome, apart from bspA (BFO_RS14480 ), revealed five more genes (BFO_RS14345 (bspB ), BFO_RS08355 , BFO_RS14330 , BFO_RS14330 , and BFO_RS14330 ) encoding putative BspA-like proteins.

    Techniques: Sequencing, Labeling

    Knockout of pseC and legC decreases biofilm formation of T. forsythia ATCC 43037 and T. forsythia UB4 cells, respectively. ( A ) Biofilm formation in T. forsythia ATCC 43037 wild-type compared to ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp . ( B ) Biofilm formation in T. forsythia UB4 wild-type compared to UB4 Δ legC and the complemented strain T. forsythia UB4 Δ legC comp . Data represent mean values ± SD of at least four independent experiments with four replicates each and were analyzed by the unpaired Student’s t-test. Asterisks indicate significant differences (** P

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: Knockout of pseC and legC decreases biofilm formation of T. forsythia ATCC 43037 and T. forsythia UB4 cells, respectively. ( A ) Biofilm formation in T. forsythia ATCC 43037 wild-type compared to ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp . ( B ) Biofilm formation in T. forsythia UB4 wild-type compared to UB4 Δ legC and the complemented strain T. forsythia UB4 Δ legC comp . Data represent mean values ± SD of at least four independent experiments with four replicates each and were analyzed by the unpaired Student’s t-test. Asterisks indicate significant differences (** P

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques: Knock-Out

    ( A ) Structures of Sia and Sia-like sugars (NulOs). Sialic acid (Neu5Ac), legionaminic acid (Leg5,7Ac 2 ) and pseudaminic acids (Pse5,7Ac 2 and Pse5Am7Gra) are shown. To note, Pse5Am7Gra is found as the terminal sugar of the S-layer glycan in T. forsythia ATCC 43037. For reference, the nine carbon atoms of Sia are numbered, and the structure of the NHAc group is shown in the boxed inset. ( B ) Schematic drawing of the structure of the S-layer O -glycan in T. forsythia ATCC 43037 (amended from Posch et al. (2011) ). This figure is available in black and white in print and in color at Glycobiology online.

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: ( A ) Structures of Sia and Sia-like sugars (NulOs). Sialic acid (Neu5Ac), legionaminic acid (Leg5,7Ac 2 ) and pseudaminic acids (Pse5,7Ac 2 and Pse5Am7Gra) are shown. To note, Pse5Am7Gra is found as the terminal sugar of the S-layer glycan in T. forsythia ATCC 43037. For reference, the nine carbon atoms of Sia are numbered, and the structure of the NHAc group is shown in the boxed inset. ( B ) Schematic drawing of the structure of the S-layer O -glycan in T. forsythia ATCC 43037 (amended from Posch et al. (2011) ). This figure is available in black and white in print and in color at Glycobiology online.

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques:

    Experimentally confirmed enzymatic steps (in green) in T. forsythia strains ATCC 43037 (left) and FDC 92A2/UB4 (right) corresponding to the CMP-Pse and CMP-Leg biosynthetic pathways as elucidated in H. pylori and C. jejuni , respectively ( Schoenhofen et al. 2006b , Schoenhofen et al. 2009 ). In T. forsythia , pathways will deviate at some point to produce the unique NulO derivatives found in our strains, such as Pse5Am7Gra. These deviations would be anticipated to occur either within the NulO biosynthetic pathway or post CMP-NulO biosynthesis. Red, blue and orange highlight enzymatic steps that introduce the stereochemical differences between the two pathways and also indicate the positions altered for both hexose intermediates and final NulO. The assignment of roman numerals to each compound is consistent with label designations found throughout the text. Subscripts “P” and “L” indicate intermediates from the CMP-Pse and CMP-Leg biosynthesis pathway, respectively. For simplicity, all sugars except for the NulOs are shown in 4 C 1 form. This figure is available in black and white in print and in color at Glycobiology online.

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: Experimentally confirmed enzymatic steps (in green) in T. forsythia strains ATCC 43037 (left) and FDC 92A2/UB4 (right) corresponding to the CMP-Pse and CMP-Leg biosynthetic pathways as elucidated in H. pylori and C. jejuni , respectively ( Schoenhofen et al. 2006b , Schoenhofen et al. 2009 ). In T. forsythia , pathways will deviate at some point to produce the unique NulO derivatives found in our strains, such as Pse5Am7Gra. These deviations would be anticipated to occur either within the NulO biosynthetic pathway or post CMP-NulO biosynthesis. Red, blue and orange highlight enzymatic steps that introduce the stereochemical differences between the two pathways and also indicate the positions altered for both hexose intermediates and final NulO. The assignment of roman numerals to each compound is consistent with label designations found throughout the text. Subscripts “P” and “L” indicate intermediates from the CMP-Pse and CMP-Leg biosynthesis pathway, respectively. For simplicity, all sugars except for the NulOs are shown in 4 C 1 form. This figure is available in black and white in print and in color at Glycobiology online.

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques: Introduce

    Phylogenetic clusters of selected microbial NulO synthase homologs. Based on the prediction of NulO sugar type by Lewis et al. (2009) , a distance-based neighbor joining tree calculated from the amino acid sequences of NulO synthase enzymes places Tanf_01240 of T. forsythia type strain ATCC 43037 in a phylogenetic clade of pseudaminic acid synthases, while BFO_1066 of strain FDC 92A2/UB4 is clustered with Leg synthases (bootstrap values shown at relevant nodes). Green denotes Leg synthases, red Neu synthases and blue Pse synthases. This figure is available in black and white in print and in color at Glycobiology online.

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: Phylogenetic clusters of selected microbial NulO synthase homologs. Based on the prediction of NulO sugar type by Lewis et al. (2009) , a distance-based neighbor joining tree calculated from the amino acid sequences of NulO synthase enzymes places Tanf_01240 of T. forsythia type strain ATCC 43037 in a phylogenetic clade of pseudaminic acid synthases, while BFO_1066 of strain FDC 92A2/UB4 is clustered with Leg synthases (bootstrap values shown at relevant nodes). Green denotes Leg synthases, red Neu synthases and blue Pse synthases. This figure is available in black and white in print and in color at Glycobiology online.

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques:

    SDS-PAGE gels (12%) of purified recombinant enzymes of the ( A ) CMP-Pse biosynthesis pathway from T. forsythia ATCC 43037 and ( B ) CMP-Leg biosynthesis pathway from T. forsythia FDC 92A2/UB4.

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: SDS-PAGE gels (12%) of purified recombinant enzymes of the ( A ) CMP-Pse biosynthesis pathway from T. forsythia ATCC 43037 and ( B ) CMP-Leg biosynthesis pathway from T. forsythia FDC 92A2/UB4.

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques: SDS Page, Purification, Recombinant

    SDS-PAGE analysis of T. forsythia parent, NulO-deficient, and complemented strains. ( A ) CBB stained SDS-PAGE (7.5% gel) of whole cell extracts from T. forsythia ATCC 43037, ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp , and ( B ) T. forsythia UB4 wild-type, UB4 Δ legC and the complemented strain UB4 Δ legC comp . For both T. forsythia ATCC 43037 Δ pseC and UB4 Δ legC , a downshift of the S-layer protein bands (TfsA and TfsB) could be observed, which was reverted in the complemented strains.

    Journal: Glycobiology

    Article Title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications

    doi: 10.1093/glycob/cww129

    Figure Lengend Snippet: SDS-PAGE analysis of T. forsythia parent, NulO-deficient, and complemented strains. ( A ) CBB stained SDS-PAGE (7.5% gel) of whole cell extracts from T. forsythia ATCC 43037, ATCC 43037 Δ pseC and the complemented strain ATCC 43037 Δ pseC comp , and ( B ) T. forsythia UB4 wild-type, UB4 Δ legC and the complemented strain UB4 Δ legC comp . For both T. forsythia ATCC 43037 Δ pseC and UB4 Δ legC , a downshift of the S-layer protein bands (TfsA and TfsB) could be observed, which was reverted in the complemented strains.

    Article Snippet: A previous bioinformatic analysis of the T. forsythia ATCC 43037 type strain genome revealed the presence of a protein O -glycosylation gene cluster encoding genes involved in sugar biosynthesis, glycan assembly and transport ( ).

    Techniques: SDS Page, Staining