Contribution of the Active Site Aspartic Acid to Catalysis in the Bacterial Neuraminidase from Micromonospora viridifaciens

JN Watson; S Newstead; V Dookhun; Garry Lindsay Taylor; A Bennett

doi:10.1016/j.febslet.2004.10.016

Contribution of the Active Site Aspartic Acid to Catalysis in the Bacterial Neuraminidase from Micromonospora viridifaciens

JN Watson, S Newstead, V Dookhun, Garry Lindsay Taylor, A Bennett

Research output: Other contribution

Abstract

A recombinant D92G mutant sialidase from Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Original language	English
Volume	577
DOIs	https://doi.org/10.1016/j.febslet.2004.10.016
Publication status	Published - 5 Nov 2004

Keywords

neuraminidase
sialidase
catalytic mechanism
mutagenesis
Micromonospora viridifaciens
VIBRIO-CHOLERAE NEURAMINIDASE
INFLUENZA-VIRUS NEURAMINIDASE
CRUZI TRANS-SIALIDASE
CRYSTAL-STRUCTURE
CLOSTRIDIUM-PERFRINGENS
GLYCOSIDASE MECHANISMS
SALMONELLA-TYPHIMURIUM
DIRECTED MUTAGENESIS
PROTEIN MODELS
REFINEMENT

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10.1016/j.febslet.2004.10.016

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title = "Contribution of the Active Site Aspartic Acid to Catalysis in the Bacterial Neuraminidase from Micromonospora viridifaciens",

abstract = "A recombinant D92G mutant sialidase from Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.",

keywords = "neuraminidase, sialidase, catalytic mechanism, mutagenesis, Micromonospora viridifaciens, VIBRIO-CHOLERAE NEURAMINIDASE, INFLUENZA-VIRUS NEURAMINIDASE, CRUZI TRANS-SIALIDASE, CRYSTAL-STRUCTURE, CLOSTRIDIUM-PERFRINGENS, GLYCOSIDASE MECHANISMS, SALMONELLA-TYPHIMURIUM, DIRECTED MUTAGENESIS, PROTEIN MODELS, REFINEMENT",

author = "JN Watson and S Newstead and V Dookhun and Taylor, {Garry Lindsay} and A Bennett",

note = "FEBS Letts",

year = "2004",

month = nov,

day = "5",

doi = "10.1016/j.febslet.2004.10.016",

language = "English",

volume = "577",

type = "Other",

}

TY - GEN

T1 - Contribution of the Active Site Aspartic Acid to Catalysis in the Bacterial Neuraminidase from Micromonospora viridifaciens

AU - Watson, JN

AU - Newstead, S

AU - Dookhun, V

AU - Taylor, Garry Lindsay

AU - Bennett, A

N1 - FEBS Letts

PY - 2004/11/5

Y1 - 2004/11/5

N2 - A recombinant D92G mutant sialidase from Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

AB - A recombinant D92G mutant sialidase from Micromonospora viridifaciens has been cloned, expressed and purified. Kinetic studies reveal that the replacement of the conserved aspartic acid with glycine results in a catalytically competent retaining sialidase that possesses significant activity against activated substrates. The contribution of this aspartate residue to the free energy of hydrolysis for natural substrates is greater than 19 kJ/mol. The three dimensional structure of the D92G mutant shows that the removal of aspartic acid 92 causes no significant re-arrangement of the active site, and that an ordered water molecule substitutes for the carboxylate group of D92. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

KW - neuraminidase

KW - sialidase

KW - catalytic mechanism

KW - mutagenesis

KW - Micromonospora viridifaciens

KW - VIBRIO-CHOLERAE NEURAMINIDASE

KW - INFLUENZA-VIRUS NEURAMINIDASE

KW - CRUZI TRANS-SIALIDASE

KW - CRYSTAL-STRUCTURE

KW - CLOSTRIDIUM-PERFRINGENS

KW - GLYCOSIDASE MECHANISMS

KW - SALMONELLA-TYPHIMURIUM

KW - DIRECTED MUTAGENESIS

KW - PROTEIN MODELS

KW - REFINEMENT

UR - http://www.scopus.com/inward/record.url?scp=7544222677&partnerID=8YFLogxK

U2 - 10.1016/j.febslet.2004.10.016

DO - 10.1016/j.febslet.2004.10.016

M3 - Other contribution

VL - 577

ER -

Contribution of the Active Site Aspartic Acid to Catalysis in the Bacterial Neuraminidase from Micromonospora viridifaciens

Abstract

Keywords

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