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Characterization of β-galactoside phosphorylases with diverging acceptor specificities

Paper ID Volume ID Publish Year Pages File Format Full-Text
17557 42677 2011 7 PDF Available
Title
Characterization of β-galactoside phosphorylases with diverging acceptor specificities
Abstract

Glycoside phosphorylases are a special group of carbohydrate-active enzymes, with characteristics in between those of glycoside hydrolases and glycosyl transferases. The phosphorylases from family GH-112 are exceptional because they employ galactose-1-phosphate instead of glucose-1-phosphate as glycosyl donor. Different acceptor specificities have been observed in this family, ranging from l-rhamnose to GlcNAc, GalNAc and a combination of the latter. Three new phosphorylases from previously unexplored branches of the phylogenetic tree of family GH-112 have now been characterized to shed more light on this divergence in acceptor specificity. The enzymes from Erysipelothrix rhusiopathiae and Streptobacillus moniliformis were found to prefer GalNAc as acceptor, while that from Anaerococcus prevotii displays similar activities on GalNAc and GlcNAc. These results confirm the correlation between the amino acid residue at position 162 and the enzyme's specificity, i.e. a threonine in the former group and a valine in the latter. However, mutagenesis of residue 162 did not allow the rational transformation of the substrate preference, as the substitution of valine by threonine in the enzyme from Bifidobacterium longum did not tighten its specificity towards GalNAc. Unexpectedly, introducing an isoleucine at position 162 increased the preference for GlcNAc as acceptor, which illustrates that the structure–function relationships in β-galactoside phosphorylases are not yet completely understood. Several other positions have also been examined by mutational analysis but true determinants of the acceptor specificity in family GH-112 could not be identified.

Keywords
Glycoside hydrolase family 112; Galacto-N-biose/lacto-N-biose I phosphorylase; β-1,4-d-Galactosyl-l-rhamnose phosphorylase; Acceptor specificity; Phylogenetic tree; Site-directed mutagenesis
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Characterization of β-galactoside phosphorylases with diverging acceptor specificities
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 49, Issue 1, 10 June 2011, Pages 59–65
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us