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Structural and enzymatic characterization of a glycoside hydrolase family 31 alpha-xylosidase from Cellvibrio japonicus involved in xyloglucan saccharification
KTH, School of Biotechnology (BIO), Glycoscience.
KTH, School of Biotechnology (BIO), Glycoscience.
KTH, School of Biotechnology (BIO), Glycoscience.
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2011 (English)In: Biochemical Journal, ISSN 0264-6021, E-ISSN 1470-8728, Vol. 436, 567-580 p.Article in journal (Refereed) Published
Abstract [en]

The desire for improved methods of biomass conversion into fuels and feedstocks has re-awakened interest in the enzymology of plant cell wall degradation. The complex polysaccharide xyloglucan is abundant in plant matter, where it may account for up to 20% of the total primary cell wall carbohydrates. Despite this, few studies have focused on xyloglucan saccharification, which requires a consortium of enzymes including endo-xyloglucanases, alpha-xylosidases, beta-galactosidases and alpha-L-fucosidases, among others. In the present paper, we show the characterization of Xy131A, a key alpha-xylosidase in xyloglucan utilization by the model Gram-negative soil saprophyte Cellvibrio japonicus. CjXy131A exhibits high regiospecificity for the hydrolysis of XGOs (xylogluco-oligosaccharides), with a particular preference for longer substrates. Crystallographic structures of both the apo enzyme and the trapped covalent 5-fluoro-beta-xylosyl-enzyme intermediate, together with docking studies with the XXXG heptasaccharide, revealed, for the first time in GH31 (glycoside hydrolase family 31), the importance of PA14 domain insert in the recognition of longer oligosaccharides by extension of the active-site pocket. The observation that CjXy131A was localized to the outer membrane provided support for a biological model of xyloglucan utilization by C. japonicas, in which XGOs generated by the action of a secreted endo-xyloglucanase are ultimately degraded in close proximity to the cell surface. Moreover, the present study diversifies the toolbox of glycosidases for the specific modification and saccharification of cell wall polymers for biotechnological applications.

Place, publisher, year, edition, pages
2011. Vol. 436, 567-580 p.
Keyword [en]
enzymology, hemicellulose, plant cell wall, saccharification, xyloglucan
National Category
Biological Sciences
URN: urn:nbn:se:kth:diva-37169DOI: 10.1042/BJ20110299ISI: 000292488500006PubMedID: 21426303ScopusID: 2-s2.0-79957714498OAI: diva2:432311

QC 20110802

Available from: 2011-08-02 Created: 2011-08-02 Last updated: 2013-08-26Bibliographically approved
In thesis
1. Strategies for the Discovery of Carbohydrate-Active Enzymes from Environmental Bacteria
Open this publication in new window or tab >>Strategies for the Discovery of Carbohydrate-Active Enzymes from Environmental Bacteria
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The focus of this thesis is a comparative study of approaches in discovery of carbohydrate-active enzymes (CAZymes). CAZymes synthesise, bind to, and degrade all the multitude of carbohydrates found in nature. As such, when aiming for sustainable methods to degrade plant biomass for the generation of biofuels, for which there is a strong drive in society, CAZymes are a natural source of environmentally friendly molecular tools.

In nature, microorganisms are the principal degraders of carbohydrates. Not only do they degrade plant matter in forests and aquatic habitats, but also break down the majority of carbohydrates ingested by animals. These symbiotic microorganisms, known as the microbiota, reside in animal digestive tracts in immense quantities, where one of the key nutrient sources is complex carbohydrates. Thus, microorganisms are a plentiful source of CAZymes, and strategies in the discovery of new enzymes from bacterial sources have been the basis for the work presented here, combined with biochemical characterisation of several enzymes.

Novel enzymatic activities for the glycoside hydrolase family 31 have been described as a result of the initial projects of the thesis. These later evolved into projects studying bacterial multi-gene systems for the partial or complete degradation of the heterogeneous plant polysaccharide xyloglucan. These systems contain, in addition to various hydrolytic CAZymes, necessary binding-, transport-, and regulatory proteins. The results presented here show, in detail, how very complex carbohydrates can efficiently be degraded by bacterial enzymes of industrial relevance.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2013. viii, 82 p.
TRITA-BIO-Report, ISSN 1654-2312 ; 2013:13
CAZyme discovery, xyloglucan, polysaccharide-utilisation locus, microbiota, α-xylosidase, GH31, transglucosidase, human gut
National Category
Biochemistry and Molecular Biology
Research subject
SRA - Molecular Bioscience
urn:nbn:se:kth:diva-126956 (URN)978-91-7501-834-8 (ISBN)
Public defence
2013-09-13, Oskar Klein Auditoriet, FR4, 10:00, Albanova Universitetscentrum, Roslagstullsbacken 21, Stockholm, 15:00 (English)

QC 20130826

Available from: 2013-08-26 Created: 2013-08-23 Last updated: 2016-01-26Bibliographically approved

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