Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.2.2.10 (PNL)
341 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The three-dimensional structure of Aspergillus niger pectin lyase B (PLB) has been determined by crystallographic techniques at a resolution of 1.7 A. The model, with all 359 amino acids and 339 water molecules, refines to a final crystallographic R factor of 16.5%. The polypeptide backbone folds into a large right-handed cylinder, termed a parallel beta helix. Loops of various sizes and conformations protrude from the central helix and probably confer function. The largest loop of 53 residues folds into a small domain consisting of three antiparallel beta strands, one turn of an alpha helix, and one turn of a 3(10) helix. By comparison with the structure of Erwinia chrysanthemi pectate lyase C (PelC), the primary sequence alignment between the pectate and pectin lyase subfamilies has been corrected and the active site region for the pectin lyases deduced. The substrate-binding site in PLB is considerably less hydrophilic than the comparable PelC region and consists of an extensive network of highly conserved Trp and His residues. The PLB structure provides an atomic explanation for the lack of a catalytic requirement for Ca2+ in the pectin lyase family, in contrast to that found in the pectate lyase enzymes. Surprisingly, however, the PLB site analogous to the Ca2+ site in PelC is filled with a positive charge provided by a conserved Arg in the pectin lyases. The significance of the finding with regard to the enzymatic mechanism is discussed.
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PMID:The tree-dimensional structure of aspergillus niger pectin lyase B at 1.7-A resolution. 944 37

Three bacterial pectate lyases, a pectin lyase from Aspergillus niger, the structures of rhamnogalacturonase A from Aspergillus aculeatus, RGase A, and the P22-phage tailspike protein, TSP, display the right-handed parallel beta-helix architecture first seen in pectate lyase. The lyases have 7 complete coils while RGase A and TSP have 11 and 12, respectively. Each coil contains three beta-strands and three turn regions named PB1, T1, PB2, T2, PB3, and T3 in their order of occurrence. The lyases have homologous sequences but RGase A and TSP do not show obvious sequence homology either to the lyases or to each other. However, the structural similarities between all these molecules are so extensive that divergence from a common ancestor is much more probable than convergence to the same fold. The region PB2-T2-PB3 is the best conserved region in the lyases and shows the clearest structural similarity. Not only is the pleating and the direction of the hydrogen bonding in the sheets conserved, but so is the unusual alphaL-conformation turn between the two sheets. However, the overall shape, the position of long loops, a conserved alpha-helix that covers the amino-terminal end of the parallel beta-helix and stacks of residues in alphaR-conformation at the start of PB1 all suggest a common ancestor. The functional similarity, that the enzymes all bind alpha-galactose containing polymers at an equivalent site involving PB1 and its two flanking turn regions, further supports divergent evolution. We suggest that the stacking of the coils and the unusual near perpendicular junction of PB2 and PB3 make the parallel beta-helix fold especially likely to maintain similar main chain conformations during divergent evolution even after all vestige of similarity in primary structure has vanished.
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PMID:Structure and evolution of parallel beta-helix proteins. 972 25

The exo-1 mutant of Neurospora crassa produced and secreted pectolytic activities when incubated in the presence of pectin-containing biological materials. This study shows that polygalacturonase, pectate lyase and pectin lyase activities were induced in media supplemented with galactose or galacturonic acid, indicating that these sugars induced the synthesis of pectinases. Pectinesterase activity was undetectable. Polygalacturonase activity was better induced by galactose than by galacturonic acid. The reverse was true for lyase activities. The inducing effect of galactose and galacturonic acid seemed to be different: (i) a mixture of galactose and galacturonic acid synergistically increased the production of pectic enzymes, as compared to that in the presence of one of these sugars; (ii) the inducing effect of galacturonic acid was partially repressed by glucose; (iii) in contrast, the inducing effect of galactose, rather than repressed, was enhanced by the presence of glucose. Altogether, these data point out to a complex mechanism of regulation of pectolytic enzymes by pectin-containing organic substances.
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PMID:Regulation of pectic enzymes from the exo-1 mutant strain of Neurospora crassa: effects of glucose, galactose, and galacturonic acid. 972 23

The methylotrophic yeast Candida boidinii S2 was found to be able to grow on pectin or polygalacturonate as a carbon source. When cells were grown on 1% (wt/vol) pectin, C. boidinii exhibited induced levels of the pectin-depolymerizing enzymes pectin methylesterase (208 mU/mg of protein), pectin lyase (673 mU/mg), pectate lyase (673 mU/mg), and polygalacturonase (3.45 U/mg) and two methanol-metabolizing peroxisomal enzymes, alcohol oxidase (0.26 U/mg) and dihydroxyacetone synthase (94 mU/mg). The numbers of peroxisomes also increased ca. two- to threefold in cells grown on these pectic compounds (3.34 and 2.76 peroxisomes/cell for cells grown on pectin and polygalacturonate, respectively) compared to the numbers in cells grown on glucose (1.29 peroxisomes/cell). The cell density obtained with pectin increased as the degree of methyl esterification of pectic compounds increased, and it decreased in strains from which genes encoding alcohol oxidase and dihydroxyacetone synthase were deleted and in a peroxisome assembly mutant. Our study showed that methanol metabolism and peroxisome assembly play important roles in the degradation of pectin, especially in the utilization of its methyl ester moieties.
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PMID:A methylotrophic pathway participates in pectin utilization by Candida boidinii. 1101 Aug 67

High-performance anion-exchange chromatography (HPAEC) coupled with a diode array detector (DAD) was used to identify and quantify oligogalacturonic acid components in pectins. Purified pectin lyase and polygalacturonase were used to generate unsaturated and saturated oligomers from pectins and sodium polygalacturonate, respectively. This method resulted in a good separation of saturated and unsaturated oligomers up to DP 13. It allowed us to follow polygalacturonase and pectate lyase depolymerisation pathways simultaneously.
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PMID:High-performance anion-exchange chromatography DAD as a tool for the identification and quantification of oligogalacturonic acids in pectin depolymerisation. 1112 36

Most structures of neutral lipases and esterases have been found to adopt the common alpha/beta hydrolase fold and contain a catalytic Ser-His-Asp triad. Some variation occurs in both the overall protein fold and in the location of the catalytic triad, and in some enzymes the role of the aspartate residue is replaced by a main-chain carbonyl oxygen atom. Here, we report the crystal structure of pectin methylesterase that has neither the common alpha/beta hydrolase fold nor the common catalytic triad. The structure of the Erwinia chrysanthemi enzyme was solved by multiple isomorphous replacement and refined at 2.4 A to a conventional crystallographic R-factor of 17.9 % (R(free) 21.1 %). This is the first structure of a pectin methylesterase and reveals the enzyme to comprise a right-handed parallel beta-helix as seen in the pectinolytic enzymes pectate lyase, pectin lyase, polygalacturonase and rhamnogalacturonase, and unlike the alpha/beta hydrolase fold of rhamnogalacturonan acetylesterase with which it shares esterase activity. Pectin methylesterase has no significant sequence similarity with any protein of known structure. Sequence conservation among the pectin methylesterases has been mapped onto the structure and reveals that the active site comprises two aspartate residues and an arginine residue. These proposed catalytic residues, located on the solvent-accessible surface of the parallel beta-helix and in a cleft formed by external loops, are at a location similar to that of the active site and substrate-binding cleft of pectate lyase. The structure of pectin methylesterase is an example of a new family of esterases.
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PMID:Three-dimensional structure of Erwinia chrysanthemi pectin methylesterase reveals a novel esterase active site. 1116 5

Colletotrichum gloeosporioides is an important pathogen of tropical and subtropical fruits. The C. gloeosporioides pelB gene was disrupted in the fungus via homologous recombination. Three independent isolates, GD-14, GD-23, and GD-29, did not produce or secrete pectate lyase B (PLB) and exhibited 25% lower pectate lyase (PL) and pectin lyase (PNL) activities and 15% higher polygalacturonase (PG) activity than the wild type. The PLB mutants exhibited no growth reduction on glucose, Na polypectate, or pectin as the sole carbon source at pH 3.8 or 6.0, except for a 15% reduction on pectin at pH 6.0. When pelB mutants were inoculated onto avocado fruits, however, a 36 to 45% reduction in estimated decay diameter was observed compared with the two controls, the wild type and undisrupted transformed isolate. In addition, these pelB mutants induced a significantly higher host phenylalanine ammonia lyase activity as well as the antifungal diene, which is indicative of higher host resistance. These results suggest that PLB is an important factor in the attack of C. gloeosporioides on avocado fruit, probably as a result of its virulence factor and role in the induction of host defense mechanisms.
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PMID:Colletotrichum gloeosporioides pelB is an important virulence factor in avocado fruit-fungus interaction. 1149 71

An improved method for assaying of the total endodepolymerase activity of pectinases has been developed. The method is based on the determination of the viscosity of a citrus pectin solution in the presence of the enzyme using an Ostwald viscometer. The depolymerizing activity of different pectinases can be detected including polygalacturonase, polymethylgalacturonase, pectin lyase, and pectate lyase. One unit of the endodepolymerase activity corresponds to the activity resulting in 50% decrease in the relative viscosity of 0.5% citrus pectin solution for 5 min at 40 degrees C and the appropriate pH. Depending on the pH-optima of the enzymes, two modifications of the method are described: 1) for acid pectinases at pH 5.0, and 2) for neutral (mildly alkaline) pectinases at pH 8.0. The modifications differed in the control and in the calculation of the activity. Six enzyme preparations were used to demonstrate the applicability of the method. The parameter used for the calculation of the enzymatic activity was directly proportional to the enzyme concentration (the dependence was linear in the range of at least 10-fold change in the enzyme concentration). The relative error of the method did not exceed 10%.
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PMID:Viscometric method for assaying of total endodepolymerase activity of pectinases. 1212 76

Eight cold-adapted, polygalacturonase-producing yeasts belonging to four species were isolated from frozen environmental samples in Iceland. They were identified as Cystofilobasidium lari-marini, Cystofilobasidium capitatum, Cryptococcus macerans and Cryptococcus aquaticus species by sequence analysis of rDNA regions. Growth behavior of the isolates was investigated. All strains could grow at 2 degrees C. Addition of glucose to pectin-containing culture medium had a repressive effect on enzyme production except for C. aquaticus, which showed increased polygalacturonase activity. Optimal temperature for enzyme production for the Cystofilobasidium strains was 14 degrees C, while that for the Cryptococcus strains was lower. Among the isolates, C. lari-marini S3B produced highest levels of enzyme activity at pH 3.2. Preliminary characterization of the polygalacturonases in the culture supernatant showed the enzyme from Cystofilobasidium strains to be optimally active at 40 degrees C and pH 5, and that from the Cryptococcus strains at 50 degrees C and pH 4. The polygalacturonase from C. macerans started to lose activity after 1 h of incubation at 40 degrees C, while that from the other strains had already lost activity at 30 degrees C. All the strains except C. aquaticus produced isoenzymes of polyglacturonase. In addition to polygalacturonase, the Cystofilobasidium strains produced pectin lyase, C. aquaticus pectin esterase, and C. macerans pectin lyase, pectate lyase and pectin esterase.
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PMID:Cold-adapted yeasts as producers of cold-active polygalacturonases. 1276 49

The effect of various parameters such as pH, agitation and aeration was studied for maximum production of pectin lyase (PL) and pectate lyase (PGL) by a novel yeast strain Debaryomyces nepalensis in bioreactor. The optimal levels of pH, aeration and agitation rate was found to be 7.0, 300rpm and 1vvm, respectively. Under these conditions, D. nepalensis produced 14,200U/L of PL and 12,000U/L of PGL corresponding to a productivity of 600U/Lh and 500U/Lh of PL and PGL, respectively. Fed-batch production was studied by feeding inducer (lemon peel), carbon source (galactose) individually and in combination at 12h of growth for enhanced production of PL and PGL. Combined feeding of inducer and carbon source at 12h was found to be the best strategy for enhanced production of PL and PGL. Under these conditions, production of PL and PGL increased to 23,300U/L and 22,400U/L, respectively which corresponded to a productivity of 728U/Lh of PL and 700U/Lh of PGL, respectively. The production was increased by 1.6- and 1.8-fold and productivity by 1.2- and 1.4-fold for PL and PGL, respectively when compared to batch culture.
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PMID:Batch and fed batch production of pectin lyase and pectate lyase by novel strain Debaryomyces nepalensis in bioreactor. 1739 59


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