Abstract
Maltooligosyltrehalose trehalohydrolase (MTHase) catalyzes the release of trehalose by cleaving the α-1,4-glucosidic linkage next to the α-1,1-linked terminal disaccharide of maltooligosyltrehalose. Computer simulation using the hydrogen bond analysis, free energy decomposition, and computational alanine scanning were employed to investigate the interaction between maltooligosyltrehalose and the enzyme. The same residues that were chosen for theoretical investigation were also studied by site-directed mutagenesis and enzyme kinetic analysis. The importance of residues determined either experimentally or computed theoretically were in good accord with each other. It was found that residues Y155, D156, and W218 of subsites -2 and -3 of the enzyme might play an important role in interacting with the ligand. The theoretically constructed structure of the enzyme-ligand complex was further validated through an ab initio quantum chemical calculation using the Gaussian09 package. The activation energy computed from this latter study was very similar to those reported in literatures for the same type of hydrolysis reactions.
Highlights
Trehalose (a-D-glucopyranosyl-a-D-glucopyranoside) is a nonreducing sugar formed from two glucose (G1) units joined by an a1,1 linkage
Domain A of Maltooligosyltrehalose trehalohydrolase (MTHase) of S. solfataricus KM1 is the catalytic domain made by a (b/a)8 barrel where both subdomains B and D are protruded from this barrel [6]
The difference in binding strength of an enzymesubstrate complex in the transition state between the wild type and mutant MTHases was estimated to examine whether a mutation would change the binding between the enzyme and substrate or not
Summary
Trehalose (a-D-glucopyranosyl-a-D-glucopyranoside) is a nonreducing sugar formed from two glucose (G1) units joined by an a1,1 linkage. The sugar has been approved as a novel food ingredient under the GRAS term in the United States and Europe [2]. Trehalose can be produced from starch by a combined enzymatic treatment using the thermophilic maltooligosyltrehalose synthase (MTSase), MTHase, and a debranching enzyme (Figure 1). Both MTSase and MTHase catalyze a side hydrolysis reaction which would decrease the yield of trehalose [3,4,5]. The structure of MTHase from Sulfolobus (S.) solfataricus KM1 is organized by three major domains namely, A, C, and E and two subdomains B and D. Domain A of MTHase of S. solfataricus KM1 is the catalytic domain made by a (b/a) barrel where both subdomains B and D are protruded from this barrel [6]
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