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Title: Structural analysis of β-glucosidase mutants derived from a hyperthermophilic tetrameric structure

Substitutive mutations that convert a tetrameric β-glucosidase into a dimeric state lead to improvement of its crystal quality. β-Glucosidase from Pyrococcus furiosus (BGLPf) is a hyperthermophilic tetrameric enzyme which can degrade cellooligosaccharides to glucose under hyperthermophilic conditions and thus holds promise for the saccharification of lignocellulosic biomass at high temperature. Prior to the production of large amounts of this enzyme, detailed information regarding the oligomeric structure of the enzyme is required. Several crystals of BGLPf have been prepared over the past ten years, but its crystal structure had not been solved until recently. In 2011, the first crystal structure of BGLPf was solved and a model was constructed at somewhat low resolution (2.35 Å). In order to obtain more detailed structural data on BGLPf, the relationship between its tetrameric structure and the quality of the crystal was re-examined. A dimeric form of BGLPf was constructed and its crystal structure was solved at a resolution of 1.70 Å using protein-engineering methods. Furthermore, using the high-resolution crystal structural data for the dimeric form, a monomeric form of BGLPf was constructed which retained the intrinsic activity of the tetrameric form. The thermostability of BGLPf is affected by its oligomeric structure. Here, the biophysicalmore » and biochemical properties of engineered dimeric and monomeric BGLPfs are reported, which are promising prototype models to apply to the saccharification reaction. Furthermore, details regarding the oligomeric structures of BGLPf and the reasons why the mutations yielded improved crystal structures are discussed.« less
Authors:
; ; ; ;  [1]
  1. National Institute of Advanced Industrial Science, 3-11-32, Kagamiyama, Higashi-Hiroshima, Hiroshima 739-0046 (Japan)
Publication Date:
OSTI Identifier:
22347781
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section D: Biological Crystallography; Journal Volume: 70; Journal Issue: Pt 3; Other Information: PMCID: PMC3949513; PMID: 24598756; PUBLISHER-ID: tz5040; OAI: oai:pubmedcentral.nih.gov:3949513; Copyright (c) Nakabayashi et al. 2014; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
Denmark
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; BIOMASS; CRYSTAL STRUCTURE; CRYSTALS; GLUCOSE; RESOLUTION; TEMPERATURE RANGE 0400-1000 K