Glucuronoxylan recognition by GH 30 xylanases: A study with enzyme and substrate variants
| Autor: | Katarína Šuchová, Peter Biely, Kristian B. R. M. Krogh, Anna Malovíková, Tine Hoff, Vladimír Puchart, Stanislav Kozmon |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Models Molecular Stereochemistry Protein Conformation 030106 microbiology Biophysics Biochemistry Polymerization Substrate Specificity 03 medical and health sciences Hydrolysis Protein structure Glucuronoxylan Glycoside hydrolase Enzyme kinetics Molecular Biology chemistry.chemical_classification Endo-1 4-beta Xylanases Glycosidic bond Kinetics 030104 developmental biology Enzyme chemistry Docking (molecular) Xylans Aspergillus niger Protein Binding |
| Zdroj: | Archives of biochemistry and biophysics. 643 |
| ISSN: | 1096-0384 |
| Popis: | XynA from Erwinia chrysanthemi (EcXyn30A), belonging to glycoside hydrolase family 30 subfamily 8, is specialized for hydrolysis of 4-O-methylglucuronoxylan (GX). Carboxyl group of 4-O-methylglucuronic acid serves as a substrate recognition element interacting ionically with positively charged Arg293 of the enzyme. We determined kinetic parameters of EcXyn30A on GX, its methyl ester (GXE) and 4-O-methylglucoxylan (GXR) and compared them with behavior of the enzyme variant in which Arg293 was replaced by Ala. The modifications of the substrate carboxyl groups resulted in several thousand-fold decrease in catalytic efficiency of EcXyn30A. In contrast, the R293A replacement reduced catalytic efficiency on GX only 18-times. The main difference was in catalytic rate (kcat) which was much lower for EcXyn30A acting on the modified substrates than for the variant which exhibited similar kcat values on all three polymers. The R293A variant cleaved GX, GXE and GXR on the second glycosidic bond from branch towards the reducing end, similarly to EcXyn30A. The R293A replacement caused 15-times decrease in specific activity on MeGlcA3Xyl4, but it did not influence low activity on linear xylooligosaccharides. Docking experiments showed that MeGlcA3Xyl4 and its esterified and reduced forms were bound to both enzymes in analogous way but with different binding energies. |
| Databáze: | OpenAIRE |
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