bamhi Search Results


bamhi  (New England Biolabs)
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New England Biolabs bamhi
Bamhi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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pt1nx vectors  (Jena Bioscience)
92
Jena Bioscience pt1nx vectors
Pt1nx Vectors, supplied by Jena Bioscience, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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spei hf  (New England Biolabs)
99
New England Biolabs spei hf
Spei Hf, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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bamhi sites  (Beyotime)
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Beyotime bamhi sites
Bamhi Sites, supplied by Beyotime, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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addgene plasmid  (Addgene inc)
93
Addgene inc addgene plasmid
Addgene Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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136546 anxa2  (Addgene inc)
90
Addgene inc 136546 anxa2
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
136546 Anxa2, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/bamhi/pm32442400-188-19-29?v=Addgene+inc
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paper addgene id  (Addgene inc)
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Addgene inc paper addgene id
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
Paper Addgene Id, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/bamhi/pm32442400-188-28-29?v=Addgene+inc
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pcdna3 1  (Addgene inc)
94
Addgene inc pcdna3 1
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
Pcdna3 1, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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restriction enzymes bamhi  (New England Biolabs)
99
New England Biolabs restriction enzymes bamhi
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
Restriction Enzymes Bamhi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/bamhi/bio_rxiv__2024__10__14__618150-163-31-38?v=New+England+Biolabs
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bamhi hf  (New England Biolabs)
99
New England Biolabs bamhi hf
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
Bamhi Hf, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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dsdna oligo  (New England Biolabs)
90
New England Biolabs dsdna oligo
Figure 2. Schematic Representation of the <t>ANXA2</t> Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.
Dsdna Oligo, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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e2f1 expression vector  (Addgene inc)
90
Addgene inc e2f1 expression vector
RNASEH2A is regulated by viral E7 via <t>E2F1</t> and its expression in association with E2F1 and PCNA in cervical and endocervical cancer and head and neck squamous cell carcinoma. (A) Read coverage maps showing the RNA expression levels of all 7 cervical cancer samples and 7 normal samples for RNASEH2A. Distributions of RNASEH2A reads (blue for normal, red for cancer) along with RNASEH2A transcript structure were visualized using the Integrative Genomics Viewer (IGV). (B) Correlation between RNASEH2A and HPV16 E7 mRNAs in 48 clinical cervical lesion samples, including 25 CIN2-CIN3 and 23 cervical cancer tissue samples. (C) siRNA-specific knockdown of HPV16 or HPV18 E7 expression in CaSki (HPV16 + ) cells or HPV18-immortalized human foreskin keratinocytes (HFK18) with respect to expression of RNASEH2A, PCNA, and NOVA1 was evaluated by Western blotting. si-NS, nonspecific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. Knockdown efficiency of E7 expression was evaluated by analysis of increased expression of p53 due to the E7 siRNA also targeting the overlapped bicistronic E6 RNA ; tubulin served as a sample loading control. (D) siRNA-specific knockdown of HPV16 or HPV18 E6 or E7 in CaSki (HPV16 + ) or HeLa (HPV18 + ) cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E6, HPV16 or HPV18 E6-specific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. (E and F) Specific-siRNA knockdown (E) or ectopic expression of E2F1 (F) in CaSki or HeLa cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E2F1, E2F1-specific siRNA; p, control vector; p-E2F1, E2F1-expression vector. * , P < 0.05; * * , P < 0.01; ** * , P < 0.001. The relative expression levels of RNASEH2A in panels D, E, and F are shown as means ± SD for each treatment group determined in duplicate from three independent experiments, with the level in the si-NS (D and E) or vector control (F) group being set to 1. (G) Increased coexpression of RNASEH2A, PCNA, and E2F1 in cervical squamous cell carcinoma, endocervical adenocarcinoma (CESC), and head and neck squamous cell carcinoma (HNSC). Data were extracted from the TCGA data sets using TCGA2STAT R package version 1.2 ( https://CRAN.R-project.org/package=TCGA2STAT ).
E2f1 Expression Vector, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Image Search Results


Figure 2. Schematic Representation of the ANXA2 Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.

Journal: Structure (London, England : 1993)

Article Title: Structure Determination of the Transactivation Domain of p53 in Complex with S100A4 Using Annexin A2 as a Crystallization Chaperone.

doi: 10.1016/j.str.2020.05.001

Figure Lengend Snippet: Figure 2. Schematic Representation of the ANXA2 Fusion Constructs Used for the Crys- tallographic Studies (A) The target protein is fused to the C-NTD (23–33) of ANXA2. Here the additional interactions between C-NTD and CTD (34–339) (red lines) stabilize the structure of ANXA2. (B) Almost the whole NTD (2–28) is removed and the target protein is fused directly to the first a helix of ANXA2 core domain (29–339) with a short linker. (C) In the present work the p53 TAD1756 (orange) was fused to the scS100A4D8 (I and II represent the subunits), which is fused to ANXA229339. Dashed lines indicate the GS linkers. Brown dots represent calcium ions.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant DNA ANXA223-339 in modified pET15b vector (pANXA223-339) This paper Addgene ID: 136543 and 136546 ANXA2 29-339 in modified pET15b vector (pANXA229-339) This paper Addgene ID: 136544 and 136545 wtMBP in in pET-MBP This paper N/A P53 TAD17-56 – scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A PDZ – ANXA223-339 in modified pET15b vector (pANXA223-339) Gogl et al., 2018 N/A wtMBP – scS100A4D8 in pET-MBP This paper N/A wtMPB – PDZ in pET-MBP Gogl et al., 2018 N/A PDZ in pET-MBP Gogl et al., 2018 N/A scS100A4D8 in modified pET15b vector (pEV) This paper N/A S100A4 in modified pET15b vector (pEV) Kiss et al., 2012 N/A P53 TAD1-60 in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide mutants in modified pGEX vector (pETARA) This paper N/A Tobacco etch virus (TEV) protease in pTH24 vector van den Berg et al., 2006 N/A Software and Algorithms Coot Emsley et al., 2010 https://www2.mrc-lmb.cam.ac.uk/personal/ Phenix Adams et al., 2010 https://www.phenix-online.org Pymol PYMOL http://www.pymol.org Phaser McCoy, 2007 http://www-structmed.cimr.cam.ac.uk/ phaser_obsolete/ XDS Kabsch, 2010 http://xds.mpimf-heidelberg.mpg.de/ Origin for ITC 5.0 OriginLab http://www.originlab.com/ Origin 8 OriginLab http://www.originlab.com/ GeneTools GeneTools http://www.syngene.com/genetools-software- download Bestsel Micsonai et al., 2015 http://bestsel.elte.hu/ Other Phenyl Sepharose 6 Fast Flow GE Healthcare #17-0973-05 HiTrap SP HP cation exchange column GE Healthcare #17115101 Jupiter 300 C5 column Phenomenex #00G-4052-E0 384-well microplates Corning #3676 Profinity IMAC resin Bio-Rad #1560131 Protino Glutathione Agarose 4B resin Macherey-Nagel #745500.10 Amylose resin NEB #E8021S HighTrap Heparin HP column GE Healthcare (Sigma) #GE17-0406-01 HiTrap Desalting column GE Healthcare (Sigma) #GE29-0486-84

Techniques: Construct

Figure 4. Crystal Screening of Different wtMBP and ANXA2 Constructs (A and B) (A) ANXA229339 and (B) PDZ-ANXA223339 crystals were formed under numerous JCSG+ (blue) and Morpheus (red) crystallization conditions at 250 mM concentration. (C–F) (C) In the case of the larger scS100A4D8-ANXA229339 construct, crystal formation significantly dropped. Crystal formation of (D) wtMBP, (E) wtMBP-PDZ, and (F) wtMBP-scS100A4D8 at 2 mM concentration is also presented. Crystal growth was monitored for a month.

Journal: Structure (London, England : 1993)

Article Title: Structure Determination of the Transactivation Domain of p53 in Complex with S100A4 Using Annexin A2 as a Crystallization Chaperone.

doi: 10.1016/j.str.2020.05.001

Figure Lengend Snippet: Figure 4. Crystal Screening of Different wtMBP and ANXA2 Constructs (A and B) (A) ANXA229339 and (B) PDZ-ANXA223339 crystals were formed under numerous JCSG+ (blue) and Morpheus (red) crystallization conditions at 250 mM concentration. (C–F) (C) In the case of the larger scS100A4D8-ANXA229339 construct, crystal formation significantly dropped. Crystal formation of (D) wtMBP, (E) wtMBP-PDZ, and (F) wtMBP-scS100A4D8 at 2 mM concentration is also presented. Crystal growth was monitored for a month.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant DNA ANXA223-339 in modified pET15b vector (pANXA223-339) This paper Addgene ID: 136543 and 136546 ANXA2 29-339 in modified pET15b vector (pANXA229-339) This paper Addgene ID: 136544 and 136545 wtMBP in in pET-MBP This paper N/A P53 TAD17-56 – scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A PDZ – ANXA223-339 in modified pET15b vector (pANXA223-339) Gogl et al., 2018 N/A wtMBP – scS100A4D8 in pET-MBP This paper N/A wtMPB – PDZ in pET-MBP Gogl et al., 2018 N/A PDZ in pET-MBP Gogl et al., 2018 N/A scS100A4D8 in modified pET15b vector (pEV) This paper N/A S100A4 in modified pET15b vector (pEV) Kiss et al., 2012 N/A P53 TAD1-60 in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide mutants in modified pGEX vector (pETARA) This paper N/A Tobacco etch virus (TEV) protease in pTH24 vector van den Berg et al., 2006 N/A Software and Algorithms Coot Emsley et al., 2010 https://www2.mrc-lmb.cam.ac.uk/personal/ Phenix Adams et al., 2010 https://www.phenix-online.org Pymol PYMOL http://www.pymol.org Phaser McCoy, 2007 http://www-structmed.cimr.cam.ac.uk/ phaser_obsolete/ XDS Kabsch, 2010 http://xds.mpimf-heidelberg.mpg.de/ Origin for ITC 5.0 OriginLab http://www.originlab.com/ Origin 8 OriginLab http://www.originlab.com/ GeneTools GeneTools http://www.syngene.com/genetools-software- download Bestsel Micsonai et al., 2015 http://bestsel.elte.hu/ Other Phenyl Sepharose 6 Fast Flow GE Healthcare #17-0973-05 HiTrap SP HP cation exchange column GE Healthcare #17115101 Jupiter 300 C5 column Phenomenex #00G-4052-E0 384-well microplates Corning #3676 Profinity IMAC resin Bio-Rad #1560131 Protino Glutathione Agarose 4B resin Macherey-Nagel #745500.10 Amylose resin NEB #E8021S HighTrap Heparin HP column GE Healthcare (Sigma) #GE17-0406-01 HiTrap Desalting column GE Healthcare (Sigma) #GE29-0486-84

Techniques: Construct, Crystallization Assay, Concentration Assay

Figure 5. Crystallization of ANXA2 Constructs and wtMBP at Different Concentrations Using the Best Conditions Found in the Screening Experiments Formation of (A) ANXA229339 (black), PDZ-ANXA223339 (gray), and scS100A4D8-ANXA229339 (horizontal lines) crystals were monitored at different protein concentrations for a month. (B) MBP crystals (vertical lines) formed only at higher than 500 mM concentration (~20.5 mg/mL). No MBP fused constructs were tested since no hit was found during screening. (C–E) (C) PDZ-ANXA223339, (D) ANXA229339, and (E) wtMBP crystals when samples were used at 250 mM (2 mM), 62.5 mM (1 mM), and 7 mM concentrations (0.5 mM) from left to right (wtMBP concentrations are in parenthesis). Small crystals are highlighted in white boxes. White bars represent 0.5 mm.

Journal: Structure (London, England : 1993)

Article Title: Structure Determination of the Transactivation Domain of p53 in Complex with S100A4 Using Annexin A2 as a Crystallization Chaperone.

doi: 10.1016/j.str.2020.05.001

Figure Lengend Snippet: Figure 5. Crystallization of ANXA2 Constructs and wtMBP at Different Concentrations Using the Best Conditions Found in the Screening Experiments Formation of (A) ANXA229339 (black), PDZ-ANXA223339 (gray), and scS100A4D8-ANXA229339 (horizontal lines) crystals were monitored at different protein concentrations for a month. (B) MBP crystals (vertical lines) formed only at higher than 500 mM concentration (~20.5 mg/mL). No MBP fused constructs were tested since no hit was found during screening. (C–E) (C) PDZ-ANXA223339, (D) ANXA229339, and (E) wtMBP crystals when samples were used at 250 mM (2 mM), 62.5 mM (1 mM), and 7 mM concentrations (0.5 mM) from left to right (wtMBP concentrations are in parenthesis). Small crystals are highlighted in white boxes. White bars represent 0.5 mm.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant DNA ANXA223-339 in modified pET15b vector (pANXA223-339) This paper Addgene ID: 136543 and 136546 ANXA2 29-339 in modified pET15b vector (pANXA229-339) This paper Addgene ID: 136544 and 136545 wtMBP in in pET-MBP This paper N/A P53 TAD17-56 – scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A PDZ – ANXA223-339 in modified pET15b vector (pANXA223-339) Gogl et al., 2018 N/A wtMBP – scS100A4D8 in pET-MBP This paper N/A wtMPB – PDZ in pET-MBP Gogl et al., 2018 N/A PDZ in pET-MBP Gogl et al., 2018 N/A scS100A4D8 in modified pET15b vector (pEV) This paper N/A S100A4 in modified pET15b vector (pEV) Kiss et al., 2012 N/A P53 TAD1-60 in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide mutants in modified pGEX vector (pETARA) This paper N/A Tobacco etch virus (TEV) protease in pTH24 vector van den Berg et al., 2006 N/A Software and Algorithms Coot Emsley et al., 2010 https://www2.mrc-lmb.cam.ac.uk/personal/ Phenix Adams et al., 2010 https://www.phenix-online.org Pymol PYMOL http://www.pymol.org Phaser McCoy, 2007 http://www-structmed.cimr.cam.ac.uk/ phaser_obsolete/ XDS Kabsch, 2010 http://xds.mpimf-heidelberg.mpg.de/ Origin for ITC 5.0 OriginLab http://www.originlab.com/ Origin 8 OriginLab http://www.originlab.com/ GeneTools GeneTools http://www.syngene.com/genetools-software- download Bestsel Micsonai et al., 2015 http://bestsel.elte.hu/ Other Phenyl Sepharose 6 Fast Flow GE Healthcare #17-0973-05 HiTrap SP HP cation exchange column GE Healthcare #17115101 Jupiter 300 C5 column Phenomenex #00G-4052-E0 384-well microplates Corning #3676 Profinity IMAC resin Bio-Rad #1560131 Protino Glutathione Agarose 4B resin Macherey-Nagel #745500.10 Amylose resin NEB #E8021S HighTrap Heparin HP column GE Healthcare (Sigma) #GE17-0406-01 HiTrap Desalting column GE Healthcare (Sigma) #GE29-0486-84

Techniques: Crystallization Assay, Construct, Concentration Assay

Figure 6. Crystal Packing of ANXA2 in Structures Found in the PDB (A–C) (A) ANXA2 in Ca2+-bound form (PDB: 1XJL, light and dark gray) (Rosengarth and Luecke, 2004) produces elongated crystal lattices (same in PDB: 5LPX, 5LQ0, 2HYW, 4X9P, and 5LQ2 structures) (Ecse´ di et al., 2017; Shao et al., 2006; Raddum et al., 2015). Structures of previously solved (B) PDZ-ANXA223339

Journal: Structure (London, England : 1993)

Article Title: Structure Determination of the Transactivation Domain of p53 in Complex with S100A4 Using Annexin A2 as a Crystallization Chaperone.

doi: 10.1016/j.str.2020.05.001

Figure Lengend Snippet: Figure 6. Crystal Packing of ANXA2 in Structures Found in the PDB (A–C) (A) ANXA2 in Ca2+-bound form (PDB: 1XJL, light and dark gray) (Rosengarth and Luecke, 2004) produces elongated crystal lattices (same in PDB: 5LPX, 5LQ0, 2HYW, 4X9P, and 5LQ2 structures) (Ecse´ di et al., 2017; Shao et al., 2006; Raddum et al., 2015). Structures of previously solved (B) PDZ-ANXA223339

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant DNA ANXA223-339 in modified pET15b vector (pANXA223-339) This paper Addgene ID: 136543 and 136546 ANXA2 29-339 in modified pET15b vector (pANXA229-339) This paper Addgene ID: 136544 and 136545 wtMBP in in pET-MBP This paper N/A P53 TAD17-56 – scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A PDZ – ANXA223-339 in modified pET15b vector (pANXA223-339) Gogl et al., 2018 N/A wtMBP – scS100A4D8 in pET-MBP This paper N/A wtMPB – PDZ in pET-MBP Gogl et al., 2018 N/A PDZ in pET-MBP Gogl et al., 2018 N/A scS100A4D8 in modified pET15b vector (pEV) This paper N/A S100A4 in modified pET15b vector (pEV) Kiss et al., 2012 N/A P53 TAD1-60 in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide mutants in modified pGEX vector (pETARA) This paper N/A Tobacco etch virus (TEV) protease in pTH24 vector van den Berg et al., 2006 N/A Software and Algorithms Coot Emsley et al., 2010 https://www2.mrc-lmb.cam.ac.uk/personal/ Phenix Adams et al., 2010 https://www.phenix-online.org Pymol PYMOL http://www.pymol.org Phaser McCoy, 2007 http://www-structmed.cimr.cam.ac.uk/ phaser_obsolete/ XDS Kabsch, 2010 http://xds.mpimf-heidelberg.mpg.de/ Origin for ITC 5.0 OriginLab http://www.originlab.com/ Origin 8 OriginLab http://www.originlab.com/ GeneTools GeneTools http://www.syngene.com/genetools-software- download Bestsel Micsonai et al., 2015 http://bestsel.elte.hu/ Other Phenyl Sepharose 6 Fast Flow GE Healthcare #17-0973-05 HiTrap SP HP cation exchange column GE Healthcare #17115101 Jupiter 300 C5 column Phenomenex #00G-4052-E0 384-well microplates Corning #3676 Profinity IMAC resin Bio-Rad #1560131 Protino Glutathione Agarose 4B resin Macherey-Nagel #745500.10 Amylose resin NEB #E8021S HighTrap Heparin HP column GE Healthcare (Sigma) #GE17-0406-01 HiTrap Desalting column GE Healthcare (Sigma) #GE29-0486-84

Techniques:

Figure 7. Crystal Contacts of Two ANXA2 Molecules (A) The aligned ANXA2 structures (PDB: 5LPX, 5LQ0, 5LPU, 1XJL, 2HYW, 4X9P, and 5LQ2) (Ecse´ di et al., 2017; Rosengarth and Luecke, 2004; Raddum et al., 2015; Shao et al., 2006) show the conservative crystal contacts. The additional segments of the crystal lattice (light gray ANXA2s are in identical position) defines the solution channel (green ellipse) formed in ANXA2 crystals. (B) S234 of light gray (molecule B) ANXA2 helps the coordination of Ca2+ (brown sphere) connecting to dark gray ANXA2 (molecule A). (C and D) (C) Besides Ca2+ coordination, residues of the nearby a helices form ionic, while (D) two tyrosines form hydrophobic interactions, further stabilizing the assembly. Residues of molecule B are underlined.

Journal: Structure (London, England : 1993)

Article Title: Structure Determination of the Transactivation Domain of p53 in Complex with S100A4 Using Annexin A2 as a Crystallization Chaperone.

doi: 10.1016/j.str.2020.05.001

Figure Lengend Snippet: Figure 7. Crystal Contacts of Two ANXA2 Molecules (A) The aligned ANXA2 structures (PDB: 5LPX, 5LQ0, 5LPU, 1XJL, 2HYW, 4X9P, and 5LQ2) (Ecse´ di et al., 2017; Rosengarth and Luecke, 2004; Raddum et al., 2015; Shao et al., 2006) show the conservative crystal contacts. The additional segments of the crystal lattice (light gray ANXA2s are in identical position) defines the solution channel (green ellipse) formed in ANXA2 crystals. (B) S234 of light gray (molecule B) ANXA2 helps the coordination of Ca2+ (brown sphere) connecting to dark gray ANXA2 (molecule A). (C and D) (C) Besides Ca2+ coordination, residues of the nearby a helices form ionic, while (D) two tyrosines form hydrophobic interactions, further stabilizing the assembly. Residues of molecule B are underlined.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Recombinant DNA ANXA223-339 in modified pET15b vector (pANXA223-339) This paper Addgene ID: 136543 and 136546 ANXA2 29-339 in modified pET15b vector (pANXA229-339) This paper Addgene ID: 136544 and 136545 wtMBP in in pET-MBP This paper N/A P53 TAD17-56 – scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A scS100A4D8 – ANXA229-339 in modified pET15b vector (pANXA229-339) This paper N/A PDZ – ANXA223-339 in modified pET15b vector (pANXA223-339) Gogl et al., 2018 N/A wtMBP – scS100A4D8 in pET-MBP This paper N/A wtMPB – PDZ in pET-MBP Gogl et al., 2018 N/A PDZ in pET-MBP Gogl et al., 2018 N/A scS100A4D8 in modified pET15b vector (pEV) This paper N/A S100A4 in modified pET15b vector (pEV) Kiss et al., 2012 N/A P53 TAD1-60 in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide in modified pGEX vector (pETARA) This paper N/A P53 TAD17-56 peptide mutants in modified pGEX vector (pETARA) This paper N/A Tobacco etch virus (TEV) protease in pTH24 vector van den Berg et al., 2006 N/A Software and Algorithms Coot Emsley et al., 2010 https://www2.mrc-lmb.cam.ac.uk/personal/ Phenix Adams et al., 2010 https://www.phenix-online.org Pymol PYMOL http://www.pymol.org Phaser McCoy, 2007 http://www-structmed.cimr.cam.ac.uk/ phaser_obsolete/ XDS Kabsch, 2010 http://xds.mpimf-heidelberg.mpg.de/ Origin for ITC 5.0 OriginLab http://www.originlab.com/ Origin 8 OriginLab http://www.originlab.com/ GeneTools GeneTools http://www.syngene.com/genetools-software- download Bestsel Micsonai et al., 2015 http://bestsel.elte.hu/ Other Phenyl Sepharose 6 Fast Flow GE Healthcare #17-0973-05 HiTrap SP HP cation exchange column GE Healthcare #17115101 Jupiter 300 C5 column Phenomenex #00G-4052-E0 384-well microplates Corning #3676 Profinity IMAC resin Bio-Rad #1560131 Protino Glutathione Agarose 4B resin Macherey-Nagel #745500.10 Amylose resin NEB #E8021S HighTrap Heparin HP column GE Healthcare (Sigma) #GE17-0406-01 HiTrap Desalting column GE Healthcare (Sigma) #GE29-0486-84

Techniques:

RNASEH2A is regulated by viral E7 via E2F1 and its expression in association with E2F1 and PCNA in cervical and endocervical cancer and head and neck squamous cell carcinoma. (A) Read coverage maps showing the RNA expression levels of all 7 cervical cancer samples and 7 normal samples for RNASEH2A. Distributions of RNASEH2A reads (blue for normal, red for cancer) along with RNASEH2A transcript structure were visualized using the Integrative Genomics Viewer (IGV). (B) Correlation between RNASEH2A and HPV16 E7 mRNAs in 48 clinical cervical lesion samples, including 25 CIN2-CIN3 and 23 cervical cancer tissue samples. (C) siRNA-specific knockdown of HPV16 or HPV18 E7 expression in CaSki (HPV16 + ) cells or HPV18-immortalized human foreskin keratinocytes (HFK18) with respect to expression of RNASEH2A, PCNA, and NOVA1 was evaluated by Western blotting. si-NS, nonspecific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. Knockdown efficiency of E7 expression was evaluated by analysis of increased expression of p53 due to the E7 siRNA also targeting the overlapped bicistronic E6 RNA ; tubulin served as a sample loading control. (D) siRNA-specific knockdown of HPV16 or HPV18 E6 or E7 in CaSki (HPV16 + ) or HeLa (HPV18 + ) cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E6, HPV16 or HPV18 E6-specific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. (E and F) Specific-siRNA knockdown (E) or ectopic expression of E2F1 (F) in CaSki or HeLa cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E2F1, E2F1-specific siRNA; p, control vector; p-E2F1, E2F1-expression vector. * , P < 0.05; * * , P < 0.01; ** * , P < 0.001. The relative expression levels of RNASEH2A in panels D, E, and F are shown as means ± SD for each treatment group determined in duplicate from three independent experiments, with the level in the si-NS (D and E) or vector control (F) group being set to 1. (G) Increased coexpression of RNASEH2A, PCNA, and E2F1 in cervical squamous cell carcinoma, endocervical adenocarcinoma (CESC), and head and neck squamous cell carcinoma (HNSC). Data were extracted from the TCGA data sets using TCGA2STAT R package version 1.2 ( https://CRAN.R-project.org/package=TCGA2STAT ).

Journal: mBio

Article Title: Genome-Wide Profiling of Cervical RNA-Binding Proteins Identifies Human Papillomavirus Regulation of RNASEH2A Expression by Viral E7 and E2F1

doi: 10.1128/mBio.02687-18

Figure Lengend Snippet: RNASEH2A is regulated by viral E7 via E2F1 and its expression in association with E2F1 and PCNA in cervical and endocervical cancer and head and neck squamous cell carcinoma. (A) Read coverage maps showing the RNA expression levels of all 7 cervical cancer samples and 7 normal samples for RNASEH2A. Distributions of RNASEH2A reads (blue for normal, red for cancer) along with RNASEH2A transcript structure were visualized using the Integrative Genomics Viewer (IGV). (B) Correlation between RNASEH2A and HPV16 E7 mRNAs in 48 clinical cervical lesion samples, including 25 CIN2-CIN3 and 23 cervical cancer tissue samples. (C) siRNA-specific knockdown of HPV16 or HPV18 E7 expression in CaSki (HPV16 + ) cells or HPV18-immortalized human foreskin keratinocytes (HFK18) with respect to expression of RNASEH2A, PCNA, and NOVA1 was evaluated by Western blotting. si-NS, nonspecific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. Knockdown efficiency of E7 expression was evaluated by analysis of increased expression of p53 due to the E7 siRNA also targeting the overlapped bicistronic E6 RNA ; tubulin served as a sample loading control. (D) siRNA-specific knockdown of HPV16 or HPV18 E6 or E7 in CaSki (HPV16 + ) or HeLa (HPV18 + ) cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E6, HPV16 or HPV18 E6-specific siRNA; si-E7, HPV16 or HPV18 E7-specific siRNA. (E and F) Specific-siRNA knockdown (E) or ectopic expression of E2F1 (F) in CaSki or HeLa cells with respect to RNASEH2A expression was evaluated by TaqMan RT-qPCR. si-NS, nonspecific siRNA; si-E2F1, E2F1-specific siRNA; p, control vector; p-E2F1, E2F1-expression vector. * , P < 0.05; * * , P < 0.01; ** * , P < 0.001. The relative expression levels of RNASEH2A in panels D, E, and F are shown as means ± SD for each treatment group determined in duplicate from three independent experiments, with the level in the si-NS (D and E) or vector control (F) group being set to 1. (G) Increased coexpression of RNASEH2A, PCNA, and E2F1 in cervical squamous cell carcinoma, endocervical adenocarcinoma (CESC), and head and neck squamous cell carcinoma (HNSC). Data were extracted from the TCGA data sets using TCGA2STAT R package version 1.2 ( https://CRAN.R-project.org/package=TCGA2STAT ).

Article Snippet: CaSki and HeLa cells were also transfected with 2 μg of an E2F1 expression vector (pRcCMV-HA-E2F1; Addgene, Watertown, MA) with X-tremeGENE HP DNA transfection reagent (Roche), and total RNA from the cells with E2F1 overexpression was analyzed for RNASEH2A expression by real-time RT-qPCR.

Techniques: Expressing, RNA Expression, Western Blot, Quantitative RT-PCR, Plasmid Preparation