Acceptor Specificity of β-N-Acetylhexosaminidase from Talaromyces flavus: A Rational Explanation

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F19%3A00519717" target="_blank" >RIV/61388971:_____/19:00519717 - isvavai.cz</a>

Výsledek na webu

<a href="https://www.mdpi.com/1422-0067/20/24/6181" target="_blank" >https://www.mdpi.com/1422-0067/20/24/6181</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3390/ijms20246181" target="_blank" >10.3390/ijms20246181</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Acceptor Specificity of β-N-Acetylhexosaminidase from Talaromyces flavus: A Rational Explanation

Popis výsledku v původním jazyce

Fungal Beta-N-acetylhexosaminidases, though hydrolytic enzymes in vivo, are useful tools in the preparation of oligosaccharides of biological interest. The Beta-N-acetylhexosaminidase from Talaromyces flavus is remarkable in terms of its synthetic potential, broad substrate specificity, and tolerance to substrate modifications. It can be heterologously produced in Pichia pastoris in a high yield. The mutation of the Tyr470 residue to histidine greatly enhances its transglycosylation capability. The aim of this work was to identify the structural requirements of this model Beta-N-acetylhexosaminidase for its transglycosylation acceptors and formulate a structure-activity relationship study. Enzymatic reactions were performed using an activated glycosyl donor, 4-nitrophenyl N-acetyl-Beta-d-glucosaminide or 4-nitrophenyl N-acetyl-Beta-d-galactosaminide, and a panel of glycosyl acceptors of varying structural features (N-acetylglucosamine, glucose, N-acetylgalactosamine, galactose, N-acetylmuramic acid, and glucuronic acid). The transglycosylation products were isolated and structurally characterized. The C-2 N-acetamido group in the acceptor molecule was found to be essential for recognition by the enzyme. The presence of the C-2 hydroxyl moiety strongly hindered the normal course of transglycosylation, yielding unique non-reducing disaccharides in a low yield. Moreover, whereas the gluco-configuration at C-4 steered the glycosylation into the Beta(1-4) position, the galacto-acceptor afforded a Beta(1-6) glycosidic linkage. The Y470H mutant enzyme was tested with acceptors based on Beta-glycosides of uronic acid and N-acetylmuramic acid. With the latter acceptor, we were able to isolate and characterize one glycosylation product in a low yield. To our knowledge, this is the first example of enzymatic glycosylation of an N-acetylmuramic acid derivative.

Název v anglickém jazyce

Acceptor Specificity of β-N-Acetylhexosaminidase from Talaromyces flavus: A Rational Explanation

Popis výsledku anglicky

Fungal Beta-N-acetylhexosaminidases, though hydrolytic enzymes in vivo, are useful tools in the preparation of oligosaccharides of biological interest. The Beta-N-acetylhexosaminidase from Talaromyces flavus is remarkable in terms of its synthetic potential, broad substrate specificity, and tolerance to substrate modifications. It can be heterologously produced in Pichia pastoris in a high yield. The mutation of the Tyr470 residue to histidine greatly enhances its transglycosylation capability. The aim of this work was to identify the structural requirements of this model Beta-N-acetylhexosaminidase for its transglycosylation acceptors and formulate a structure-activity relationship study. Enzymatic reactions were performed using an activated glycosyl donor, 4-nitrophenyl N-acetyl-Beta-d-glucosaminide or 4-nitrophenyl N-acetyl-Beta-d-galactosaminide, and a panel of glycosyl acceptors of varying structural features (N-acetylglucosamine, glucose, N-acetylgalactosamine, galactose, N-acetylmuramic acid, and glucuronic acid). The transglycosylation products were isolated and structurally characterized. The C-2 N-acetamido group in the acceptor molecule was found to be essential for recognition by the enzyme. The presence of the C-2 hydroxyl moiety strongly hindered the normal course of transglycosylation, yielding unique non-reducing disaccharides in a low yield. Moreover, whereas the gluco-configuration at C-4 steered the glycosylation into the Beta(1-4) position, the galacto-acceptor afforded a Beta(1-6) glycosidic linkage. The Y470H mutant enzyme was tested with acceptors based on Beta-glycosides of uronic acid and N-acetylmuramic acid. With the latter acceptor, we were able to isolate and characterize one glycosylation product in a low yield. To our knowledge, this is the first example of enzymatic glycosylation of an N-acetylmuramic acid derivative.

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

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2019

Kód důvěrnosti údajů

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

Název periodika

International Journal of Molecular Sciences

ISSN

1422-0067

e-ISSN

—

Svazek periodika

20

Číslo periodika v rámci svazku

24

Stát vydavatele periodika

CH - Švýcarská konfederace

Počet stran výsledku

17

Strana od-do

6181

Kód UT WoS článku

000506840100075

EID výsledku v databázi Scopus

2-s2.0-85076363105