Please use this identifier to cite or link to this item: https://doi.org/10.1016/j.jmb.2008.02.041
Title: Crystal Structure of the Polyextremophilic α-Amylase AmyB from Halothermothrix orenii: Details of a Productive Enzyme-Substrate Complex and an N Domain with a Role in Binding Raw Starch
Authors: Tan, T.-C.
Mijts, B.N.
Swaminathan, K. 
Patel, B.K.C.
Divne, C.
Keywords: α-amylase
enzyme-substrate complex
methionine interaction
N domain
polyextremophilic
Issue Date: 9-May-2008
Citation: Tan, T.-C., Mijts, B.N., Swaminathan, K., Patel, B.K.C., Divne, C. (2008-05-09). Crystal Structure of the Polyextremophilic α-Amylase AmyB from Halothermothrix orenii: Details of a Productive Enzyme-Substrate Complex and an N Domain with a Role in Binding Raw Starch. Journal of Molecular Biology 378 (4) : 850-868. ScholarBank@NUS Repository. https://doi.org/10.1016/j.jmb.2008.02.041
Abstract: The gene for a membrane-bound, halophilic, and thermostable α-amylase, AmyB, from Halothermothrix orenii was cloned and sequenced. The crystal structure shows that, in addition to the typical domain organization of family 13 glycoside hydrolases, AmyB carries an additional N-terminal domain (N domain) that forms a large groove-the N-C groove-some 30 Å away from the active site. The structure of AmyB with the inhibitor acarbose at 1.35 Å resolution shows that a nonasaccharide has been synthesized through successive transglycosylation reactions of acarbose. Unexpectedly, in a complex of wild-type AmyB with α-cyclodextrin and maltoheptaose at 2.2 Å resolution, a maltotetraose molecule is bound in subsites - 1 to + 3, spanning the cleavage point at - 1/+ 1, with the - 1 glucosyl residue present as a 2So skew boat. This wild-type AmyB complex was obtained in the presence of a large excess of substrate, a condition under which it is possible to capture Michaelis complexes, which may explain the observed binding across - 1/+ 1 and ring distortion. We observe three methionine side chains that serve as "binding platforms" for glucosyl rings in AmyB, a seemingly rare occurrence in carbohydrate-binding proteins. The structures and results from the biochemical characterization of AmyB and AmyB lacking the N domain show that the N domain increases binding of the enzyme to raw starch. Furthermore, theoretical modeling suggests that the N-C groove can accommodate, spatially and chemically, large substrates such as A-starch. © 2008 Elsevier Ltd. All rights reserved.
Source Title: Journal of Molecular Biology
URI: http://scholarbank.nus.edu.sg/handle/10635/100369
ISSN: 00222836
DOI: 10.1016/j.jmb.2008.02.041
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