The beta-N-Acetylhexosaminidase in the Synthesis of Bioactive Glycans: Protein and Reaction Engineering

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F19%3A00503890" target="_blank" >RIV/61388971:_____/19:00503890 - isvavai.cz</a>

Výsledek na webu

<a href="https://www.mdpi.com/1420-3049/24/3/599" target="_blank" >https://www.mdpi.com/1420-3049/24/3/599</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3390/molecules24030599" target="_blank" >10.3390/molecules24030599</a>

Jazyk výsledku

angličtina

Název v původním jazyce

The beta-N-Acetylhexosaminidase in the Synthesis of Bioactive Glycans: Protein and Reaction Engineering

Popis výsledku v původním jazyce

N-Acetylhexosamine oligosaccharides terminated with GalNAc act as selective ligands of galectin-3, a biomedically important human lectin. Their synthesis can be accomplished by beta-N-acetylhexosaminidases (EC 3.2.1.52). Advantageously, these enzymes tolerate the presence of functional groups in the substrate molecule, such as the thiourea linker useful for covalent conjugation of glycans to a multivalent carrier, affording glyconjugates. beta-N-Acetylhexosaminidases exhibit activity towards both N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) moieties. A point mutation of active-site amino acid Tyr into other amino acid residues, especially Phe, His, and Asn, has previously been shown to strongly suppress the hydrolytic activity of beta-N-acetylhexosaminidases, creating enzymatic synthetic engines. In the present work, we demonstrate that Tyr470 is an important mutation hotspot for altering the ratio of GlcNAcase/GalNAcase activity, resulting in mutant enzymes with varying affinity to GlcNAc/GalNAc substrates. The enzyme selectivity may additionally be manipulated by altering the reaction medium upon changing pH or adding selected organic co-solvents. As a result, we are able to fine-tune the beta-N-acetylhexosaminidase affinity and selectivity, resulting in a high-yield production of the functionalized GalNAc beta 4GlcNAc disaccharide, a selective ligand of galectin-3.

Název v anglickém jazyce

The beta-N-Acetylhexosaminidase in the Synthesis of Bioactive Glycans: Protein and Reaction Engineering

Popis výsledku anglicky

N-Acetylhexosamine oligosaccharides terminated with GalNAc act as selective ligands of galectin-3, a biomedically important human lectin. Their synthesis can be accomplished by beta-N-acetylhexosaminidases (EC 3.2.1.52). Advantageously, these enzymes tolerate the presence of functional groups in the substrate molecule, such as the thiourea linker useful for covalent conjugation of glycans to a multivalent carrier, affording glyconjugates. beta-N-Acetylhexosaminidases exhibit activity towards both N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) moieties. A point mutation of active-site amino acid Tyr into other amino acid residues, especially Phe, His, and Asn, has previously been shown to strongly suppress the hydrolytic activity of beta-N-acetylhexosaminidases, creating enzymatic synthetic engines. In the present work, we demonstrate that Tyr470 is an important mutation hotspot for altering the ratio of GlcNAcase/GalNAcase activity, resulting in mutant enzymes with varying affinity to GlcNAc/GalNAc substrates. The enzyme selectivity may additionally be manipulated by altering the reaction medium upon changing pH or adding selected organic co-solvents. As a result, we are able to fine-tune the beta-N-acetylhexosaminidase affinity and selectivity, resulting in a high-yield production of the functionalized GalNAc beta 4GlcNAc disaccharide, a selective ligand of galectin-3.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

<a href="/cs/project/LTC18038" target="_blank" >LTC18038: Multivalentní glyko-biomateriály s potenciálem v regenerační medicíně</a><br>

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

Molecules

ISSN

1420-3049

e-ISSN

—

Svazek periodika

24

Číslo periodika v rámci svazku

3

Stát vydavatele periodika

CH - Švýcarská konfederace

Počet stran výsledku

14

Strana od-do

599

Kód UT WoS článku

000458934000223

EID výsledku v databázi Scopus

2-s2.0-85061365096