Abstract
In the present work we suggest an efficient method, using the whole time course of the reaction, whereby parameters kcat, Km and product KI for the hydrolysis of a p-nitrophenyl glycoside by an exo-acting glycoside hydrolase can be estimated in a single experiment. Its applicability was demonstrated for three retaining exo-glycoside hydrolases, β-xylosidase from Aspergillus awamori, β-galactosidase from Penicillium sp. and α-galactosidase from Thermotoga maritima (TmGalA). During the analysis of the reaction course catalyzed by the TmGalA enzyme we had observed that a non-enzymatic process, mutarotation of the liberated α-d-galactose, affected the reaction significantly.
Original language | English |
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Pages (from-to) | 43-49 |
Number of pages | 7 |
Journal | Carbohydrate Research |
Volume | 412 |
DOIs | |
Publication status | Published - 14 Aug 2015 |
MoE publication type | A1 Journal article-refereed |
Funding
The work was supported by the Russian Foundation for Basic Research (project No. 12-08-00813-a ), a personal scholarship from the Swedish Institute to A.B., and personal fellowship of the Government of Leningrad District, St. Petersburg, Russia to G.R. NMR experiments were carried out using scientific equipment of the Center of Shared Usage “The analytical center of nano- and biotechnologies of Saint Petersburg State Polytechnical University” financially supported by the Ministry of Education and Science of the Russian Federation .
Keywords
- Integrated kinetics
- Mutarotation
- Retaining glycoside hydrolase