Hypertransglycosylating Variants of the GH20 beta-N-Acetylhexosaminidase for the Synthesis of Chitooligomers

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F22%3A00558832" target="_blank" >RIV/61388971:_____/22:00558832 - isvavai.cz</a>

Výsledek na webu

<a href="https://onlinelibrary.wiley.com/doi/epdf/10.1002/adsc.202200046" target="_blank" >https://onlinelibrary.wiley.com/doi/epdf/10.1002/adsc.202200046</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1002/adsc.202200046" target="_blank" >10.1002/adsc.202200046</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Hypertransglycosylating Variants of the GH20 beta-N-Acetylhexosaminidase for the Synthesis of Chitooligomers

Popis výsledku v původním jazyce

Fungal beta-N-acetylhexosaminidases of the CAZy family 20 of glycoside hydrolases are well-established tools for the enzymatic synthesis of a wide variety of natural and modified oligosaccharides and glycoconjugates. In order to increase their synthetic efficiency, the beta-N-acetylhexosaminidase from Aspergillus oryzae (AoHex) was employed as a model enzyme for enzyme engineering aiming at shifting the reaction course from hydrolysis toward transglycosylation. Specifically, nine mutant variants of AoHex were designed by molecular modeling based on its crystal structure and molecular dynamics simulations. The selected mutation hotspots included the tyrosine residue at the active site, which stabilizes the transition state of the reaction, and two residues at the aglycone-binding site, which were replaced by tryptophan residues to increase the hydrophobicity of this subsite. Besides the individual mutants, combined double-mutant variants were also prepared and characterized. As a result, eight out of the studied new AoHex variants had transglycosidase activity, with V306W/Y445N AoHex being a superior transglycosidase with a transglycosylation-to-hydrolysis ratio greater than 110, which is entirely unique among the hypertransglycosylating glycosidase mutants including the GH20 beta-N-acetylhexosaminidases.

Název v anglickém jazyce

Hypertransglycosylating Variants of the GH20 beta-N-Acetylhexosaminidase for the Synthesis of Chitooligomers

Popis výsledku anglicky

Fungal beta-N-acetylhexosaminidases of the CAZy family 20 of glycoside hydrolases are well-established tools for the enzymatic synthesis of a wide variety of natural and modified oligosaccharides and glycoconjugates. In order to increase their synthetic efficiency, the beta-N-acetylhexosaminidase from Aspergillus oryzae (AoHex) was employed as a model enzyme for enzyme engineering aiming at shifting the reaction course from hydrolysis toward transglycosylation. Specifically, nine mutant variants of AoHex were designed by molecular modeling based on its crystal structure and molecular dynamics simulations. The selected mutation hotspots included the tyrosine residue at the active site, which stabilizes the transition state of the reaction, and two residues at the aglycone-binding site, which were replaced by tryptophan residues to increase the hydrophobicity of this subsite. Besides the individual mutants, combined double-mutant variants were also prepared and characterized. As a result, eight out of the studied new AoHex variants had transglycosidase activity, with V306W/Y445N AoHex being a superior transglycosidase with a transglycosylation-to-hydrolysis ratio greater than 110, which is entirely unique among the hypertransglycosylating glycosidase mutants including the GH20 beta-N-acetylhexosaminidases.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2022

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Advanced Synthesis & Catalysis

ISSN

1615-4150

e-ISSN

1615-4169

Svazek periodika

364

Číslo periodika v rámci svazku

12

Stát vydavatele periodika

DE - Spolková republika Německo

Počet stran výsledku

14

Strana od-do

2009-2022

Kód UT WoS článku

000796769700001

EID výsledku v databázi Scopus

2-s2.0-85130509136