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Function of second glucan binding site including tyrosines 54 and 101 in Thermus aquaticus amylomaltase

Paper ID Volume ID Publish Year Pages File Format Full-Text
22333 43269 2007 7 PDF Available
Title
Function of second glucan binding site including tyrosines 54 and 101 in Thermus aquaticus amylomaltase
Abstract

Amylomaltase from Thermus aquaticus catalyzes three types of transglycosylation reaction, as well as a weak hydrolytic reaction of α-1,4 glucan. From our previous study [Fujii et al., Appl. Environ. Microbiol., 71, 5823–5827 (2005)], tyrosine 54 (Y54) was identified as an amino acid controlling the reaction specificity of this enzyme. Since Y54 is not located around the active site but in the proposed second glucan binding site that is 14 Å away from catalytic residues, the functions of Y54 and the second glucan binding site are of great interest. In this study, we introduced mutations into another tyrosine (Y101) in the second glucan binding site. The obtained mutated enzymes were subjected to all four types of enzyme assay and the effects of mutations on the reaction specificities of these enzymes were comprehensively investigated. These studies indicated that the amino acid substitution at Y54 or Y101 for removing their aromatic side chain increases cyclization activity (intra-molecular transglycosylation reaction) but decreases disproportionation, coupling and hydrolytic activities (inter-molecular reactions). The superimposition of the reported structures of the enzyme with and without substrate analog revealed the occurrence of a conformational change in which a donor binding site becomes open. From lines of evidence, we conclude that the binding of glucan substrate to the second glucan binding site through an interaction with the aromatic side chains of Y54 and Y101 is a trigger for the enzyme to take a completely active conformation for all four types of activity, but prevents the cyclization reaction to occur since the flexibility of the glucan is restricted by such binding.

Keywords
amylomaltase; Thermus aquaticus; transglycosylation; cycloamylose
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Function of second glucan binding site including tyrosines 54 and 101 in Thermus aquaticus amylomaltase
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 103, Issue 2, February 2007, Pages 167–173
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