Journal: International Journal of Molecular Sciences

Article Title: Metabolic Response of the Yeast Candida utilis During Enrichment in Selenium

doi: 10.3390/ijms21155287

Figure Lengend Snippet: Effect of selenium on the growth of Candida utilis yeast cells.

Article Snippet: The authors found that C. utilis CCTCC M 209298 could accumulate approximately 1010 μg Se/g after bioreactor culture in medium supplemented with 15 mg Se/L.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Metabolic Response of the Yeast Candida utilis During Enrichment in Selenium

doi: 10.3390/ijms21155287

Figure Lengend Snippet: Parameters characterizing the growth Candida utilis ATCC 9950 during culturing in the control YPD medium and experimental media enriched in selenium.

Article Snippet: The authors found that C. utilis CCTCC M 209298 could accumulate approximately 1010 μg Se/g after bioreactor culture in medium supplemented with 15 mg Se/L.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: Metabolic Response of the Yeast Candida utilis During Enrichment in Selenium

doi: 10.3390/ijms21155287

Figure Lengend Snippet: Results of selenium speciation analysis of Candida yeast.

Article Snippet: The authors found that C. utilis CCTCC M 209298 could accumulate approximately 1010 μg Se/g after bioreactor culture in medium supplemented with 15 mg Se/L.

Techniques:

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Purification of PIMT from C. utilis . Data represented are the average of three sets of experimentations.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Purification, Activity Assay, High Performance Liquid Chromatography

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Activity of Purified Native Protein l -Isoaspartyl Methyltransferase from C. utilis toward Different l -Isoaspartate- containing Peptide and Protein Substrates. The endogenous activity in the absence of methyl-accepting substrate is subtracted. Enzyme concentrations used for the assays was 35 µM. All reactions were done in triplicates. Data represented in the table is the mean from three independent experimental states with±range.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Activity Assay, Purification, Concentration Assay

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Kinetic Rate Constants of Purified Native Protein l -Isoaspartyl Methyltransferase from C. utilis . Methyltransferase reactions were performed as described under Materials and Methods in triplicate using varying concentrations of the substrates Iso-Asp DSIP (0–0.1 mM) and [ 3 H]AdoMet (0–10 µM). For the reactions with variable peptide concentrations, AdoMet was used at a concentration of 40 µM whereas for variable AdoMet concentrations, Iso-Asp DSIP was used as a substrate at 0.3 mM. 10 µl enzyme protein was used for all experiments. The kinetic parameters of the enzyme were calculated by fitting the data to the Michaelis–Menten equation using the Origin (Version 8.5) software. K i was determined by fitting the data into Lineweaver–Burke plots for the different concentrations of AdoHcy used. Values represent mean±SD.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Purification, Concentration Assay, Software

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Structural characteristics of the native CuPCMT . (A) Amino acid sequence of the CuPCMT was determined by MALDI TOF MS/MS analysis. The AdoMet binding sites within the sequence were determined by analyzing the sequence with online tool PSIPRED and are underlined in red; (B). Circular dichroism spectra of the purified PCMT. The X -axis represents the wavelength in nm and Y -axis represents molar ellipticity ( θ ). Deconvolution of the CD data by CDNN software gave percentages of the α-helix, β-sheet and random coil. The predicted and experimentally determined percentages are listed in ; (C). To validate the amino acid sequence for structural characteristics, it was analyzed by CD search tool from NCBI. The output image denotes the conserved domains detected within the amino acid sequence; D. Phylogenetic comparisons between PCMT from Candida and eleven homologs from different representative organisms by Clustal Omega tool gave a Phylogenetic tree in the form of a rooted cladogram. The abbreviations used are: THEMA: Thermotoga maritima ; HELPY: Helicobacter pylori ; CANUT: Candida utilis ; SCHPO: Schizosaccharomyces pombe ; NEUCR: Neurospora crassa ; TRIAE: Triticum aestivum ; ARATH: Arabidopsis thaliana ; CAEEL: Caenorhabditis elegans ; DROME: Drosophila melanogaster ; BOVIN: Bovine.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Sequencing, Tandem Mass Spectroscopy, Binding Assay, Circular Dichroism, Purification, Software

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: In vivo regulation of CuPCMT in C. utilis . (A) C. utilis cells were grown up to early stationary phase and were incubated either with 0–2 mM AdOx or with 0–2 mM LiCl for 24 h. Post incubation cells were harvested, washed, lysed and checked for PCMT activity. Y -axis denotes PCMT activity in terms of pmoles of methyl groups transferred/min/ml enzyme. X -axis denotes both the concentration of AdOx and/or LiCl in mM. (B) ROS generation profile of cells treated by different concentrations of AdOx. The X -axis denotes the concentrations of AdOx used and the Y -axis denotes the percentage of cells showing H 2 DCFDA fluorescence; (C) ROS generation profile of cells treated by different concentrations of LiCl. The X -axis denotes the concentrations of LiCl used and the Y -axis denotes the percentage of cells showing H 2 DCFDA fluorescence; (D) Semi-quantitative RT-PCR analysis of CuPCMT transcript concentrations in cells incubated in different LiCl concentrations is exhibited in the top panel. β-actin transcript expression of the same sets of cells are designated in the lower panel; (E) Early stationary phase Candida cells were treated with 0.8 mM AdOx for 24 h followed by oxidative ( ), pH ( ), hyperosmotic ( ) and heat ( ) stress. Isoapartate levels of treated and untreated cells were worked out in terms of pmoles of isoAsp/pmole total cellular proteins. (F) Early stationary phase Candida cells were incubated in presence or absence of 1.2 mM LiCl for 24 h after which the cells were subjected to either oxidative ( ), pH ( ), hyperosmotic ( ) and heat ( ) stress. Intracellular isoAsp levels from control and treated sets were determined in terms of pmoles of isoAsp/pmole total cellular proteins. Data presented is from two independent experimentation sets. The horizontal bars indicate the average (Mean), and the error bars represent standard deviation (S.D.).

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: In Vivo, Incubation, Activity Assay, Concentration Assay, Fluorescence, Quantitative RT-PCR, Expressing, Control, Standard Deviation

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Role of CuPCMT in cell survival under physiological stress . Control set represents early stationary phase C. utilis cells reconstituted in fresh YPD medium without AdOx or LiCl and not subjected to any stress condition. Untreated sets represent C. utilis cells reconstituted in media without AdOx or LiCl but subjected to the same stress condition as the treated sets. AdOx set represents C. utilis cells reconstituted in YPD medium containing 0.8 mM AdOx and subjected to stress condition. LiCl set represents C. utilis cells reconstituted in YPD medium containing 1 mM LiCl and subjected to stress conditions. Candida cells were subjected to oxidative stress by addition of 30 mM H 2 O 2 for 200 min following 24 h incubation in either 0.8 mM AdOx or in 1 mM LiCl. Aliquots were drawn every 50 min. (A) Cell viability was assayed colorimetrically with MTT treatment and (B) cell death was monitored by flow cytometry analysis. Candida utilis cells were subjected to pH stress for with 0.1 mM Hydrochloric acid for 6 h following 24 h incubation in either 0.8 mM AdOx or 1 mM LiCl. Aliquots were drawn every 1 h. (C) Cell viability was assessed colorimetrically by MTT assay and (D) cell death by staining with Annexin V-FITC/PI followed by FACS analysis. C. utilis cells were subjected to hyperosmotic stress for 40 mins with 1.5 M NaCl following 24 h incubation in either 0.8 mM AdOx or in 1 mM LiCl. Aliquots were drawn at 10 min intervals from both treated and untreated sets and tested for (E) cell viability by MTT assay as well as (F) cell survival by flow cytometry after staining with Annexin V-FITC conjugate/PI. Candida utilis cells were subjected to heat stress at 42 °C for 40 min following 24 h incubation in either 0.8 mM AdOx or 1 mM LiCl. Aliquots were drawn every 10 min. (G) Cell viability was assessed colorimetrically by MTT assay and (H) cell death by staining with Annexin V-FITC/PI followed by FACS analysis.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Control, Incubation, Flow Cytometry, MTT Assay, Staining

Journal: Biochemistry and Biophysics Reports

Article Title: Enzymatic attributes of an l -isoaspartyl methyltransferase from Candida utilis and its role in cell survival

doi: 10.1016/j.bbrep.2015.08.015

Figure Lengend Snippet: Identification of possible substrates of PIMT in AdOx treated C. utilis cells . Coomassie-stained spots corresponding to major methyl acceptors ( Fig. 10 ) were subjected to peptide mass fingerprinting by MALDI-TOF. The MS data was matched with the Uniprot protein database against all fungal proteins as well as with NCBI non-redundant protein database.

Article Snippet: The wild type C. utilis strain was obtained from National Chemical Laboratories, Pune, India (Cat. No. NCIM Y500).

Techniques: Peptide Mass Fingerprinting, Glycoproteomics, Modification