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

Garcia-Oliva, Cecilia ; Hoyos, Pilar ; Petrásková, Lucie ; Kulik, Natalia ; Pelantová, Helena ; Cabanillas, Alfredo H ; Rumbero, Ángel ; Křen, Vladimír ; Hernáiz, María J ; Bojarová, Pavla

International journal of molecular sciences, 2019-12, Vol.20 (24), p.6181 [Periódico revisado por pares]

Switzerland: MDPI AG

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Título:
Acceptor Specificity of β- N -Acetylhexosaminidase from Talaromyces flavus : A Rational Explanation
Autor: Garcia-Oliva, Cecilia ; Hoyos, Pilar ; Petrásková, Lucie ; Kulik, Natalia ; Pelantová, Helena ; Cabanillas, Alfredo H ; Rumbero, Ángel ; Křen, Vladimír ; Hernáiz, María J ; Bojarová, Pavla
Assuntos: Acids ; beta-N-Acetylhexosaminidases - chemistry ; beta-N-Acetylhexosaminidases - metabolism ; Chromatography ; Data acquisition ; Disaccharides ; Enzymes ; Galactose ; Glucose ; Glycosides ; Glycosides - metabolism ; Glycosylation ; Histidine ; Kinetics ; Models, Molecular ; Mutation ; N-Acetylgalactosamine ; N-Acetylglucosamine ; Oligosaccharides ; Oligosaccharides - metabolism ; Protein Conformation ; Spectrum analysis ; Structure-Activity Relationship ; Substrate Specificity ; Substrates ; Talaromyces ; Talaromyces - enzymology ; Uronic acid
É parte de: International journal of molecular sciences, 2019-12, Vol.20 (24), p.6181
Notas: ObjectType-Article-1
SourceType-Scholarly Journals-1
ObjectType-Feature-2
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Descrição: Fungal β- -acetylhexosaminidases, though hydrolytic enzymes in vivo, are useful tools in the preparation of oligosaccharides of biological interest. The β- -acetylhexosaminidase from is remarkable in terms of its synthetic potential, broad substrate specificity, and tolerance to substrate modifications. It can be heterologously produced in 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 β- -acetylhexosaminidase for its transglycosylation acceptors and formulate a structure-activity relationship study. Enzymatic reactions were performed using an activated glycosyl donor, 4-nitrophenyl -acetyl-β-d-glucosaminide or 4-nitrophenyl -acetyl-β-d-galactosaminide, and a panel of glycosyl acceptors of varying structural features ( -acetylglucosamine, glucose, -acetylgalactosamine, galactose, -acetylmuramic acid, and glucuronic acid). The transglycosylation products were isolated and structurally characterized. The C-2 -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 -configuration at C-4 steered the glycosylation into the β(1-4) position, the -acceptor afforded a β(1-6) glycosidic linkage. The Y470H mutant enzyme was tested with acceptors based on β-glycosides of uronic acid and -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 -acetylmuramic acid derivative. In order to explain these findings and predict enzyme behavior, a modeling study was accomplished that correlated with the acquired experimental data.
Editor: Switzerland: MDPI AG
Idioma: Inglês

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

Garcia-Oliva, Cecilia ; Hoyos, Pilar ; Petrásková, Lucie ; Kulik, Natalia ; Pelantová, Helena ; Cabanillas, Alfredo H ; Rumbero, Ángel ; Křen, Vladimír ; Hernáiz, María J ; Bojarová, Pavla

Switzerland: MDPI AG

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