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Hypertransglycosylating Variants of the GH20 beta-N-Acetylhexosaminidase for the Synthesis of Chitooligomers
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SYSNO ASEP 0558832 Druh ASEP J - Článek v odborném periodiku Zařazení RIV J - Článek v odborném periodiku Poddruh J Článek ve WOS Název Hypertransglycosylating Variants of the GH20 beta-N-Acetylhexosaminidase for the Synthesis of Chitooligomers Tvůrce(i) Mészáros, Zuzana (MBU-M)
Petrásková, Lucie (MBU-M) ORCID
Kulik, Natalia (MBU-M) ORCID
Pelantová, Helena (MBU-M) ORCID, RID
Bojarová, Pavla (MBU-M) ORCID
Křen, Vladimír (MBU-M) RID, ORCID
Slámová, Kristýna (MBU-M) RID, ORCIDZdroj.dok. Advanced Synthesis & Catalysis. - : Wiley - ISSN 1615-4150
Roč. 364, č. 12 (2022), s. 2009-2022Poč.str. 14 s. Jazyk dok. eng - angličtina Země vyd. DE - Německo Klíč. slova chitinase ; protein ; expression ; tryptophan ; mutants ; yasara ; biotransformations ; enzymes ; glycosylation ; oligosaccharides ; protein engineering ; protein models Vědní obor RIV CE - Biochemie Obor OECD Biochemistry and molecular biology CEP GA20-00477S GA ČR - Grantová agentura ČR LTC19035 GA MŠMT - Ministerstvo školství, mládeže a tělovýchovy LM2018131 GA MŠMT - Ministerstvo školství, mládeže a tělovýchovy Výzkumná infrastruktura e-INFRA CZ - 90140 - CESNET, zájmové sdružení právnických osob Způsob publikování Omezený přístup Institucionální podpora MBU-M - RVO:61388971 UT WOS 000796769700001 EID SCOPUS 85130509136 DOI 10.1002/adsc.202200046 Anotace 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. Pracoviště Mikrobiologický ústav Kontakt Eliška Spurná, eliska.spurna@biomed.cas.cz, Tel.: 241 062 231 Rok sběru 2023 Elektronická adresa https://onlinelibrary.wiley.com/doi/epdf/10.1002/adsc.202200046
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