The Experts below are selected from a list of 282 Experts worldwide ranked by ideXlab platform
Dinesh Yadav - One of the best experts on this subject based on the ideXlab platform.
-
Molecular Cloning and Structural Insights into Pectin Lyase proteins from different strains of Fusarium
Current Proteomics, 2020Co-Authors: Sangeeta Yadav, Vinay Kumar Singh, Gautam Anand, Dinesh YadavAbstract:: Pectin Lyaseis an industrially important enzymeof Pectinase group that degrade Pectin polymers forming 4,5-unsaturated oligogalacturonides. Several fugal Pectin Lyase genes predominately from Aspergillus and Penicillium genera have been reported in the literature. Five Pectin Lyase genes were cloned from FusariumoxysporumMTCC1755, F.monoliforme var. subglutinansMTCC2015, FusariumavneceumMTCC10572, and FusariumsolaniMTCC3004 using PCR approach. Pectin Lyase genes and proteins were subjected to homology search, multiple sequence alignment, motif search, physio-chemical characterization, phylogenetic tree construction, 3D structure prediction and molecular docking. Many conserved amino acids were found at several positions in all the Pectin Lyase proteins. Phylogenetic analysis of these proteins alongwith other Pectinases revealed two major clusters representing members of Lyases and hydrolases. In-silico characterization revealed Pectin Lyase proteins to be highly stable owing to the presence of disulfide bonds in their structure. Molecular weight and pI of these proteins were in the range 14.4 to 25.1 kDa and 4.47-9.39 respectively. Pectin Lyase proteins from different Fusariumstrains were very much similar in their structural features and biochemical properties which might be due to their similarity on the primary sequence. Docking studies revealed that electrostatic forces, vander Waal and hydrogen bonds are the major interacting forces between the ligands and the enzyme. This might be accountable for comparatively higher and better activity of Pectin Lyase against galacturonic acid as compared to α-D-galactopyranuronic acid, galactofuranuronicacid and galactopyranuronate. Aspartate, tyrosine and tryptophan residues in the active site of the enzyme are responsible for ligand binding.
-
Molecular Biology, Genomics and Bioinformatics Insights into Fungal Pectin Lyase: An overview
Applications of Biotechnology for Sustainable Development, 2017Co-Authors: S. Yadav, Pramod Kumar Yadav, Gautam Anand, Amit Dubey, Aiman Tanveer, R. Dwivedi, Dinesh YadavAbstract:Pectinase represents an industrially important group of enzymes, comprising Pectin Lyases (PNL), pectate Lyases (PL), polygalacturonases (PG), and Pectin methylesterases (PME) as a well-studied member with diverse applications. Among Pectinases, Pectin Lyase occupies unique positions based on its reaction mechanisms. Pectin Lyase is associated with degradation of Pectin polymer directly by β-elimination, and 4, 5-unsaturated oligogalacturonide is the product formed. In case of other Pectinases, sequential degradation of Pectin molecule occurs. Further, its relevance in fruit juice clarification is mainly because of two reasons. First, they are known to degrade Pectin without altering the ester group responsible for the specific aroma of the juice and second, the highly toxic methanol is not generated by the process of degradation of Pectin-by-Pectin Lyases. The applications of Pectin Lyases in retting processes have recently been elucidated. There exist substantial reports of fungal Pectin Lyases especially from Aspergillus and Penicillium species, with sole emphasis on production optimization, purification, and biochemical characterizations. As far as molecular biology of Pectin Lyases is concerned, few studies have been performed especially on Aspergillus species and molecular cloning and expression of relevant Pectin Lyases genes have been reported. Efforts have been made to decipher the factors influencing Pectinases gene regulation, though not extensively. Molecular cloning of Pectin Lyase gene from diverse sources, its manipulation by using directed evolution techniques, and searching for novel sources by utilizing metagenomic-based approach are some of the recent developments in Pectin Lyase research, which is a subject matter for the present review.
-
Purification, characterization and retting of Crotolaria juncea fibres by an alkaline Pectin Lyase from Fusarium oxysporum MTCC 1755.
3 Biotech, 2017Co-Authors: S. Yadav, Gautam Anand, R. Dwivedi, S. K. Maurya, Dinesh YadavAbstract:Using solid-state fermentation, production of an industrially important Pectin Lyase from a fungal strain Fusarium oxysporum MTCC 1755 was attempted, which was further subjected to purification and characterization. The enzyme was purified by three steps, namely ammonium sulfate fractionation, cation-exchange chromatography on CM cellulose followed by gel filtration chromatography using Sephadex G-100 column. A 16-fold purification with 31.2% yield and 3.2 U/mg specific activity was achieved. The optimum pH of the purified enzyme was 9.0 and stability ranged from pH 5.0–7.0 for 24 h. Optimum temperature of purified enzyme was found to be 40 °C while temperature stability ranged from 10 to 50 °C for 30 min. The K m and k cat of the enzyme was 1.75 mg/ml and 83.3 s−1, respectively. The purified enzyme was found to be highly stimulated by Ca2+ ions while sugars like mannitol and sorbitol, and salts like NaCl and CaCl2 enhanced the thermostability. The purified Pectin Lyase was found suitable for retting of Crotolaria juncea fiber.
-
Purification and characterization of a highly alkaline Pectin Lyase from Fusarium lateritum MTCC 8794
Biologia, 2017Co-Authors: Sangeeta Yadav, Gautam Anand, S. K. Maurya, Ritika Dwivedi, Dinesh YadavAbstract:A highly alkaline Pectin Lyase (PNL) produced by Fusarium lateritum MTCC 8794 using solid-state fermentation was purified and biochemically characterized. The enzyme was purified to homogeneity by comparatively simple method involving ammonium sulfate precipitation and cation exchange chromatography resulting in a final purification fold of 5.5 with specific activity of 1.9 U/mg and yield of 3.42%. The SDS-PAGE of the purified enzyme revealed a single protein band of approximately 16 kDa. The pH optimum was found to be 10.0, while the enzyme was stable in the pH range 6.0–10.0. The optimum temperature of the purified PNL was 40 °C, the enzyme being stable upto 50 °C for 30 min. The K _m value calculated by Michaelis-Menten curve was found to be 0.79 mg/mL, while V _max and k_cat of the purified enzyme were found to be 0.57 international unit and 41.6 s^−1, respectively. The enzyme showed inhibition by most of the divalent cations at 1 mM concentration.
-
PCR Amplification, Sequencing, and In Silico Characterization of Pectin Lyase Genes from Aspergillus flavus NIICC8142
Microbial Diversity and Biotechnology in Food Security, 2014Co-Authors: Amit Dubey, Sangeeta Yadav, Gautam Anand, Dinesh YadavAbstract:The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven Pectin Lyase (pnl) genes, which were used to design gene-specific primers and these genes were amplified through polymerase chain reaction (PCR) from five strains of A. flavus, namely A. flavus MTCC 7589, A. flavus MTCC 10938, A. flavus MTCC 8836, A. flavus NIICC 8142 and A. flavus NIICC 8147. All the seven predicted pnl genes were amplified from the genomic DNA of A. flavus NIICC 8142, while six pnl genes were from A. flavus MTCC8837 and five pnl genes from the remaining strains, respectively. A total of five pnl gene sequences of A. flavus NIICC 8142 designated as Afpnl-1, Afpnl-2, Afpnl-3, Afpnl-4, and Afpnl-5 were submitted to GenBank and assigned accession numbers JQ735890 to JQ735894, respectively. These sequences were subjected to in silico characterization for homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The homology search revealed their identity to the predicted Pectin Lyase genes of A. flavus NRRL 3357, and multiple sequence alignment of these five genes showed various conserved residues. The phylogenetic tree revealed two distinct clusters and four subclusters with five Pectin Lyase genes of A. flavus occupying distinct position among the PCR-amplified Pectin Lyase genes from different fungi. The presence of unique pec_Lyase C domain was observed among these sequences.
Sangeeta Yadav - One of the best experts on this subject based on the ideXlab platform.
-
Molecular Cloning and Structural Insights into Pectin Lyase proteins from different strains of Fusarium
Current Proteomics, 2020Co-Authors: Sangeeta Yadav, Vinay Kumar Singh, Gautam Anand, Dinesh YadavAbstract:: Pectin Lyaseis an industrially important enzymeof Pectinase group that degrade Pectin polymers forming 4,5-unsaturated oligogalacturonides. Several fugal Pectin Lyase genes predominately from Aspergillus and Penicillium genera have been reported in the literature. Five Pectin Lyase genes were cloned from FusariumoxysporumMTCC1755, F.monoliforme var. subglutinansMTCC2015, FusariumavneceumMTCC10572, and FusariumsolaniMTCC3004 using PCR approach. Pectin Lyase genes and proteins were subjected to homology search, multiple sequence alignment, motif search, physio-chemical characterization, phylogenetic tree construction, 3D structure prediction and molecular docking. Many conserved amino acids were found at several positions in all the Pectin Lyase proteins. Phylogenetic analysis of these proteins alongwith other Pectinases revealed two major clusters representing members of Lyases and hydrolases. In-silico characterization revealed Pectin Lyase proteins to be highly stable owing to the presence of disulfide bonds in their structure. Molecular weight and pI of these proteins were in the range 14.4 to 25.1 kDa and 4.47-9.39 respectively. Pectin Lyase proteins from different Fusariumstrains were very much similar in their structural features and biochemical properties which might be due to their similarity on the primary sequence. Docking studies revealed that electrostatic forces, vander Waal and hydrogen bonds are the major interacting forces between the ligands and the enzyme. This might be accountable for comparatively higher and better activity of Pectin Lyase against galacturonic acid as compared to α-D-galactopyranuronic acid, galactofuranuronicacid and galactopyranuronate. Aspartate, tyrosine and tryptophan residues in the active site of the enzyme are responsible for ligand binding.
-
Purification and characterization of a highly alkaline Pectin Lyase from Fusarium lateritum MTCC 8794
Biologia, 2017Co-Authors: Sangeeta Yadav, Gautam Anand, S. K. Maurya, Ritika Dwivedi, Dinesh YadavAbstract:A highly alkaline Pectin Lyase (PNL) produced by Fusarium lateritum MTCC 8794 using solid-state fermentation was purified and biochemically characterized. The enzyme was purified to homogeneity by comparatively simple method involving ammonium sulfate precipitation and cation exchange chromatography resulting in a final purification fold of 5.5 with specific activity of 1.9 U/mg and yield of 3.42%. The SDS-PAGE of the purified enzyme revealed a single protein band of approximately 16 kDa. The pH optimum was found to be 10.0, while the enzyme was stable in the pH range 6.0–10.0. The optimum temperature of the purified PNL was 40 °C, the enzyme being stable upto 50 °C for 30 min. The K _m value calculated by Michaelis-Menten curve was found to be 0.79 mg/mL, while V _max and k_cat of the purified enzyme were found to be 0.57 international unit and 41.6 s^−1, respectively. The enzyme showed inhibition by most of the divalent cations at 1 mM concentration.
-
PCR Amplification, Sequencing, and In Silico Characterization of Pectin Lyase Genes from Aspergillus flavus NIICC8142
Microbial Diversity and Biotechnology in Food Security, 2014Co-Authors: Amit Dubey, Sangeeta Yadav, Gautam Anand, Dinesh YadavAbstract:The genomic sequences of Aspergillus flavus NRRL 3357 revealed seven Pectin Lyase (pnl) genes, which were used to design gene-specific primers and these genes were amplified through polymerase chain reaction (PCR) from five strains of A. flavus, namely A. flavus MTCC 7589, A. flavus MTCC 10938, A. flavus MTCC 8836, A. flavus NIICC 8142 and A. flavus NIICC 8147. All the seven predicted pnl genes were amplified from the genomic DNA of A. flavus NIICC 8142, while six pnl genes were from A. flavus MTCC8837 and five pnl genes from the remaining strains, respectively. A total of five pnl gene sequences of A. flavus NIICC 8142 designated as Afpnl-1, Afpnl-2, Afpnl-3, Afpnl-4, and Afpnl-5 were submitted to GenBank and assigned accession numbers JQ735890 to JQ735894, respectively. These sequences were subjected to in silico characterization for homology search, multiple sequence alignment, phylogenetic tree construction, and motif analysis. The homology search revealed their identity to the predicted Pectin Lyase genes of A. flavus NRRL 3357, and multiple sequence alignment of these five genes showed various conserved residues. The phylogenetic tree revealed two distinct clusters and four subclusters with five Pectin Lyase genes of A. flavus occupying distinct position among the PCR-amplified Pectin Lyase genes from different fungi. The presence of unique pec_Lyase C domain was observed among these sequences.
-
Purification and biochemical characterization of an alkaline Pectin Lyase from Fusarium decemcellulare MTCC 2079 suitable for Crotalaria juncea fiber retting.
Journal of basic microbiology, 2013Co-Authors: Sangeeta Yadav, Gautam Anand, Amit Kumar Dubey, Reetesh Kumar, Dinesh YadavAbstract:An extracellular Pectin Lyase secreted by Fusarium decemcellulare MTCC 2079 under solid state fermentation condition has been purified to electrophoretic homogeniety by using ammonium sulfate fractionation, carboxymethyl cellulose and gel filtration (Sephadex G-100) column chromatographies. The purified enzyme showed single protein band corresponding to molecular mass 45 ± 01 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme had maximum activity at pH 9.0 and showed maximum stability in the pH range of 9.0-12.0. The optimum temperature of the purified enzyme was 50 °C and it showed maximum stability upto 40 °C. The energy of activation for the thermal denaturation (Ea ) was 59.06 kJ mol(-1) K(-1). The Km and kcat values using citrus Pectin as the substrate were 0.125 mg ml(-1) and 72.9 s(-1) in 100 mM sodium carbonate buffer pH 9.0 at 50 °C. The biophysical studies on Pectin Lyase showed that its secondary structure belongs to α + β class of protein with comparatively less of β-sheets. Purified Pectin Lyase showed efficient retting of Crotolaria juncea fibers.
-
Characterization of a neutral Pectin Lyase produced by Oidiodendron echinulatum MTCC 1356 in solid state fermentation.
Journal of basic microbiology, 2012Co-Authors: Sangeeta Yadav, Gautam Anand, Amit Dubey, Dinesh YadavAbstract:A neutral Pectin Lyase produced by a new fungal strain Oidiodendron echinulatum MTCC 1356 under solid state fermentation using wheat bran as agro waste has been studied. The enzyme was purified by ammonium sulphate precipitation (30-60%), DEAE anion exchange and Sephadex G-100 column chromatographies. The SDS-PAGE and native PAGE revealed two bands of sizes 42 and 47 kDa. The enzyme was purified 37 fold with specific activity of 4.5 U/mg and 2.25% yield. The K(m) and V(max) values determined using citrus Pectin were 1.2 mg/ml and 0.36 IU/min respectively. The pH and temperature optima were pH 7.0 and 50 °C, respectively. The pH stability was around 5.0 for 24 h at 20 °C. The purified enzyme retained maximum activity for 30 min upto 50 °C. The activation energy for thermal denaturation of the purified enzyme was found to be 60.0 kJ/Mol. The effects of various metal ions and protein inhibitors on enzyme activity have revealed total inhibition of the enzyme activity in the presence of Ag(+) and Cu(+) and KMnO(4) at 1 mM. The neutral Pectin Lyase showed retting of Crotalaria juncea fibre in the presence of EDTA.
K. D. S. Yadav - One of the best experts on this subject based on the ideXlab platform.
-
In silico analysis of Pectin Lyase and Pectinase sequences
Biochemistry. Biokhimiia, 2009Co-Authors: Pramod Kumar Yadav, Vinay Kumar Singh, Sarita Yadav, K. D. S. Yadav, Dinesh YadavAbstract:A total of 48 full-length protein sequences of Pectin Lyases from different source organisms available in NCBI were subjected to multiple sequence alignment, domain analysis, and phylogenetic tree construction. A phylogenetic tree constructed on the basis of the protein sequences revealed two distinct clusters representing Pectin Lyases from bacterial and fungal sources. Similarly, the multiple accessions of different source organisms representing bacterial and fungal Pectin Lyases also formed distinct clusters, showing sequence level homology. The sequence level similarities among different groups of Pectinase enzymes, viz. Pectin Lyase, pectate Lyase, polygalacturonase, and Pectin esterase, were also analyzed by subjecting a single protein sequence from each group with common source organism to tree construction. Four distinct clusters representing different groups of Pectinases with common source organisms were observed, indicating the existing sequence level similarity among them. Multiple sequence alignment of Pectin Lyase protein sequence of different source organisms along with Pectinases with common source organisms revealed a conserved region, indicating homology at sequence level. A conserved domain Pec_Lyase_C was frequently observed in the protein sequences of Pectin Lyases and pectate Lyases, while Glyco_hydro_28 domains and Pectate Lyase-like beta-helix clan domain are frequently observed in polygalacturonases and Pectin esterases, respectively. The signature amino acid sequence of 41 amino acids, i.e. TYDNAGVLPITVNSNKSLIGEGSKGVIKGKGLRIVSGAKNI, related with the Pec_Lyase_C is frequently observed in Pectin Lyase protein sequences and might be related with the structure and enzymatic function.
-
Purification and characterization of Pectin Lyase secreted by Penicillium citrinum.
Biochemistry. Biokhimiia, 2009Co-Authors: Sarita Yadav, Pramod Kumar Yadav, Dinesh Yadav, K. D. S. YadavAbstract:The importance of various parameters such as sugarcane juice concentration, pH of the medium, and effects of different solid supports for maximum secretion of Pectin Lyase from Penicillium citrinum MTCC 8897 has been studied. The enzyme was purified to homogeneity by Sephadex G-100 and DEAE-cellulose chromatography. The molecular mass determined by SDS-PAGE was 31 kDa. The K m and k cat values were found to be 1 mg/ml and 76 sec−1, respectively. The optimum pH of the purified Pectin Lyase was 9.0, though it retains activity in the pH 9.0–12.0 range when exposed for 24 h. The optimum temperature was 50°C, and the Pectin Lyase was found to be completely stable up to 40°C when exposed for 1 h. The purified Pectin Lyase was found efficient in retting of Linum usitatissimum, Cannabis sativa, and Crotalaria juncea.
-
Pectin Lyase: A review
Process Biochemistry, 2009Co-Authors: Sangeeta Yadav, Pramod Kumar Yadav, Dinesh Yadav, K. D. S. YadavAbstract:Abstract Pectin Lyase acts on the pectic substances that occur as structural polysaccharides in the middle lamella and primary cell walls of higher plants. This enzyme has potential applications in food, paper and textile industries. Since new applications of this enzyme are emerging there is a scientific need to explore the important aspects of the enzyme specifically the catalytic efficiency and possible sources. Though the research work on Pectin Lyase has been done for the last six decades but there is no exclusive review on Pectin Lyase so far available in the literature. This review tries to fill this gap by providing all relevant information exclusively for Pectin Lyase. The topics covered in this review are a brief description of the substrate Pectin, enzymes related to Pectin Lyase, assay procedures, sources, purification and characterization, structural aspects, substrate specificity, molecular biology, biotechnological applications and future prospects of Pectin Lyases.
-
Purification and characterization of an alkaline Pectin Lyase from Aspergillus flavus
Process Biochemistry, 2008Co-Authors: Sangeeta Yadav, Pramod Kumar Yadav, Dinesh Yadav, K. D. S. YadavAbstract:Abstract An alkaline Pectin Lyase secreted by Aspergillus flavus MTCC 7589 was purified to electrophoretic homogeneity using ammonium sulphate fractionation, anion exchange chromatography on DEAE cellulose and gel filtration chromatography on sephadex G-100. The pH and temperature optima of the enzyme were found to be 8.0 and 50 °C. The enzyme was found to be stable for 24 h in the pH range 4.0–10.0. The enzyme does not loose activity up to 50 °C if exposed for 1 h. Addition of ammonium sulphate in the range of 0.1–2.0 M increased the thermostability of the enzyme, 0.6 and 1.8 M of ammonium sulphate providing complete stability at 60 and 70 °C respectively. The values of Km and kcat of the enzyme were 0.59 mg/ml and 52.2 s−1 respectively. The molecular weight was found to be 38 ± 01 kDa. The purified enzyme showed efficacy in retting of Crotalaria juncea fibers.
Kazuo Izaki - One of the best experts on this subject based on the ideXlab platform.
-
Production, Purification, and Properties of a Pectin Lyase from Pseudomonas marginalis N6301
Bioscience Biotechnology and Biochemistry, 1995Co-Authors: Naoki Nikaidou, Yoshiyuki Kamio, Takatoshi Naganuma, Kazuo IzakiAbstract:Pseudomonas marginalis N6301 produced Pectin Lyase (EC 4.2.2.10) in the medium with cell lysis when the culture was treated with mitomycin C. We purified the enzyme by carboxymethylcellulose, hydroxylapatite, and gel-filtration column chromatographies. The enzyme had a molecular weight of 34,000 by SDS polyacrylamide gel electrophoresis and was mostly stable around pH 6.5. The optimum pH and temperature for the enzyme activity were 8.0 and 30°C, respectively. The activity was inhibited severely by 2 mM N-ethylmaleimide, maleic anhydride, and p-chloromercuriphenylsulfonic acid. Further, the Pectin Lyase produced in a medium containing glycerol was purified. The molecular weight of the enzyme was identical to that of the enzyme produced in the presence of mitomycin C.
-
Expression of a Pectin Lyase Gene in Escherichia coli from Pseudomonas marginalis N6301
Bioscience biotechnology and biochemistry, 1994Co-Authors: Naoki Nikaidou, Yoshiyuki Kamio, Kazuo IzakiAbstract:We constructed deletion mutant clones of a Pectin Lyase gene, and measured their Pectin Lyase activities in Escherichia coli. Pectin Lyase activities were detected only in a recA+ strain but not in a recA- strain of E. coli. We also cloned and sequenced recA from Pseudomonas marginalis N6301. The recA from P. marginalis N6301 can complement recA- to form recA+ in the phenotype of E. coli. Highly conserved sequences of recA are observed among E. coli, P. fluorescens, and P. marginalis. From these results, we presume that recA is required for the expression of the Pectin Lyase gene in P. marginalis N6301.
-
Analysis of Promoter Region of the Pectin Lyase Gene from Erwinia carotovora Er
Bioscience biotechnology and biochemistry, 1994Co-Authors: Hiroki Ohnishi, Naoki Nikaidou, Yoshiyuki Kamio, Kazuo IzakiAbstract:A Pectin Lyase defective mutant was constructed from Erwinia carotovora Er by transposon Tn5 insertion mutagenesis to analyze the promoter region of the pnl gene, which had been cloned. The promoter of pnl is between –140 and – 74 upstream of the structural gene of pnl and appears not to be regulated by Lex A.
Pramod Kumar Yadav - One of the best experts on this subject based on the ideXlab platform.
-
Molecular Biology, Genomics and Bioinformatics Insights into Fungal Pectin Lyase: An overview
Applications of Biotechnology for Sustainable Development, 2017Co-Authors: S. Yadav, Pramod Kumar Yadav, Gautam Anand, Amit Dubey, Aiman Tanveer, R. Dwivedi, Dinesh YadavAbstract:Pectinase represents an industrially important group of enzymes, comprising Pectin Lyases (PNL), pectate Lyases (PL), polygalacturonases (PG), and Pectin methylesterases (PME) as a well-studied member with diverse applications. Among Pectinases, Pectin Lyase occupies unique positions based on its reaction mechanisms. Pectin Lyase is associated with degradation of Pectin polymer directly by β-elimination, and 4, 5-unsaturated oligogalacturonide is the product formed. In case of other Pectinases, sequential degradation of Pectin molecule occurs. Further, its relevance in fruit juice clarification is mainly because of two reasons. First, they are known to degrade Pectin without altering the ester group responsible for the specific aroma of the juice and second, the highly toxic methanol is not generated by the process of degradation of Pectin-by-Pectin Lyases. The applications of Pectin Lyases in retting processes have recently been elucidated. There exist substantial reports of fungal Pectin Lyases especially from Aspergillus and Penicillium species, with sole emphasis on production optimization, purification, and biochemical characterizations. As far as molecular biology of Pectin Lyases is concerned, few studies have been performed especially on Aspergillus species and molecular cloning and expression of relevant Pectin Lyases genes have been reported. Efforts have been made to decipher the factors influencing Pectinases gene regulation, though not extensively. Molecular cloning of Pectin Lyase gene from diverse sources, its manipulation by using directed evolution techniques, and searching for novel sources by utilizing metagenomic-based approach are some of the recent developments in Pectin Lyase research, which is a subject matter for the present review.
-
Purification and Characterization of Pectin Lyase Produced by Aspergillus terricola and its Application in Retting of Natural Fibers
Applied Biochemistry and Biotechnology, 2009Co-Authors: Sangeeta Yadav, Pramod Kumar Yadav, Dinesh Yadav, Kapil Deo Singh YadavAbstract:An indigenously isolated fungal strain identified as Aspergillus terricola with assigned fungal strain number MTCC 7588 has been used as source for Pectin Lyase production. The extracellular Pectin Lyase was purified to homogeneity from the culture filtrate of A. terricola by ion exchange and gel filtration chromatography. The determined molecular weight was 35 ± 01 kDa. The K _m and k _cat (turnover) values of the purified enzyme at 37 °C using citrus Pectin as the substrate were found to be 1.0 mg/ml and 110.0 s^−1, respectively. The pH and temperature optima of the enzyme were 8.0 and 50 °C, respectively. The retting ability of the purified Pectin Lyase for natural fibers viz. Cannabis sativa and Linum usitatissimum has been demonstrated for the first time.
-
In silico analysis of Pectin Lyase and Pectinase sequences
Biochemistry. Biokhimiia, 2009Co-Authors: Pramod Kumar Yadav, Vinay Kumar Singh, Sarita Yadav, K. D. S. Yadav, Dinesh YadavAbstract:A total of 48 full-length protein sequences of Pectin Lyases from different source organisms available in NCBI were subjected to multiple sequence alignment, domain analysis, and phylogenetic tree construction. A phylogenetic tree constructed on the basis of the protein sequences revealed two distinct clusters representing Pectin Lyases from bacterial and fungal sources. Similarly, the multiple accessions of different source organisms representing bacterial and fungal Pectin Lyases also formed distinct clusters, showing sequence level homology. The sequence level similarities among different groups of Pectinase enzymes, viz. Pectin Lyase, pectate Lyase, polygalacturonase, and Pectin esterase, were also analyzed by subjecting a single protein sequence from each group with common source organism to tree construction. Four distinct clusters representing different groups of Pectinases with common source organisms were observed, indicating the existing sequence level similarity among them. Multiple sequence alignment of Pectin Lyase protein sequence of different source organisms along with Pectinases with common source organisms revealed a conserved region, indicating homology at sequence level. A conserved domain Pec_Lyase_C was frequently observed in the protein sequences of Pectin Lyases and pectate Lyases, while Glyco_hydro_28 domains and Pectate Lyase-like beta-helix clan domain are frequently observed in polygalacturonases and Pectin esterases, respectively. The signature amino acid sequence of 41 amino acids, i.e. TYDNAGVLPITVNSNKSLIGEGSKGVIKGKGLRIVSGAKNI, related with the Pec_Lyase_C is frequently observed in Pectin Lyase protein sequences and might be related with the structure and enzymatic function.
-
Purification and characterization of Pectin Lyase secreted by Penicillium citrinum.
Biochemistry. Biokhimiia, 2009Co-Authors: Sarita Yadav, Pramod Kumar Yadav, Dinesh Yadav, K. D. S. YadavAbstract:The importance of various parameters such as sugarcane juice concentration, pH of the medium, and effects of different solid supports for maximum secretion of Pectin Lyase from Penicillium citrinum MTCC 8897 has been studied. The enzyme was purified to homogeneity by Sephadex G-100 and DEAE-cellulose chromatography. The molecular mass determined by SDS-PAGE was 31 kDa. The K m and k cat values were found to be 1 mg/ml and 76 sec−1, respectively. The optimum pH of the purified Pectin Lyase was 9.0, though it retains activity in the pH 9.0–12.0 range when exposed for 24 h. The optimum temperature was 50°C, and the Pectin Lyase was found to be completely stable up to 40°C when exposed for 1 h. The purified Pectin Lyase was found efficient in retting of Linum usitatissimum, Cannabis sativa, and Crotalaria juncea.
-
Pectin Lyase: A review
Process Biochemistry, 2009Co-Authors: Sangeeta Yadav, Pramod Kumar Yadav, Dinesh Yadav, K. D. S. YadavAbstract:Abstract Pectin Lyase acts on the pectic substances that occur as structural polysaccharides in the middle lamella and primary cell walls of higher plants. This enzyme has potential applications in food, paper and textile industries. Since new applications of this enzyme are emerging there is a scientific need to explore the important aspects of the enzyme specifically the catalytic efficiency and possible sources. Though the research work on Pectin Lyase has been done for the last six decades but there is no exclusive review on Pectin Lyase so far available in the literature. This review tries to fill this gap by providing all relevant information exclusively for Pectin Lyase. The topics covered in this review are a brief description of the substrate Pectin, enzymes related to Pectin Lyase, assay procedures, sources, purification and characterization, structural aspects, substrate specificity, molecular biology, biotechnological applications and future prospects of Pectin Lyases.