Functional analysis of glycoside hydrolase family 8 xylanases shows narrow but distinct substrate specificities and biotechnological potential
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The potential of glycoside hydrolase family (GH) 8 xylanases in biotechnological applications is virtually unexplored. Therefore, the substrate preference and hydrolysis product profiles of two GH8 xylanases were evaluated to investigate their activities and substrate specificities. A GH8 xylanase from an uncultured bacterium (rXyn8) shows endo action but very selectively releases xylotriose from its substrates. It has a higher activity than the Pseudoalteromonas haloplanktis GH8 endo-xylanase (PhXyl) on xylononaose and smaller xylo-oligosaccharides. PhXyl preferably degrades xylan substrates with a high degree of polymerization. It is sterically more hindered by arabinose substituents than rXyn8, producing larger end hydrolysis products. The specificities of rXyn8 and PhXyl differ completely from these of the previously described GH8 xylanases from Bifidobacterium adolescentis (BaRexA) and Bacillus halodurans (BhRex). As reducing-end xylose-releasing exo-oligoxylanases, they selectively release xylose from the reducing end of small xylo-oligosaccharides. The findings of this study show that GH8 xylanases have a narrow substrate specificity, but also one that strongly varies between family members and is distinct from that of GH10 and GH11 xylanases. Structural comparison of rXyn8, PhXyl, BaRexA, and BhRex showed that subtle amino acid changes in the glycon as well as the aglycon subsites probably form the basis of the observed differences between GH8 xylanases. GH8 xylanases, therefore, are an interesting group of enzymes, with potential towards engineering and applications.
KeywordXylanase Glycoside hydrolase family 8 Substrate specificity Arabinoxylan Xylo-oligosaccharides
The authors like to thank Dr. Charles C. Lee of the Agricultural Research Service (Albany, CA, USA) for donation of plasmid DNA from rXyn8 and Prof. Anna Kulminskaya from the Russian Academy of Science (St. Petersburg, Russia) for the labeled XOS. Financial support for the SBO IMPAXOS project by the ‘Instituut voor de aanmoediging van Innovatie door Wetenschap en Technologie in Vlaanderen’ (I.W.T., Brussels, Belgium), for the post-doctoral fellowship of E.D. by the ‘Fonds voor Wetenschappelijk Onderzoek-Vlaanderen’ (F.W.O, Brussels, Belgium) and from the Research Fund K.U.Leuven (GOA/03/10 and IDO/03/005) are gratefully acknowledged. S.V.S. thanks the Biomedical Sciences Group (K.U. Leuven) for a start-up grant. The study is part of the Methusalem program ‘Food for the Future’ at the K.U. Leuven.
- Brennan Y, Callen WN, Christoffersen L, Dupree P, Goubet F, Healey S, Hernandez M, Keller M, Li K, Palackal N, Sittenfeld A, Tamayo G, Wells S, Hazlewood GP, Mathur EJ, Short JM, Robertson DE, Steer BA (2004) Unusual microbial xylanases from insect guts. Appl Environ Microbiol 70:3609–3617CrossRefGoogle Scholar
- Broekaert WF, Courtin CA, Verbeke K, Van de Wiele T, Verstraete W, Delcour JA (2010) Prebiotic and other health-related effects of cereal-derived arabinoxylans, arabinoxylan-oligosaccharides and xylo-oligosaccharides. Crit Rev Food Sci Nutr, Accepted for publicationGoogle Scholar
- Butt MS, Tahir-Nadeem M, Ahmad Z, Sultan MT (2008) Xylanases and their applications in baking industry. Food Technol Biotechnol 46:22–31Google Scholar
- De Vos D, Collins T, Nerinckx W, Savvides SN, Claeyssens M, Gerday C, Feller G, Van Beeumen J (2006) Oligosaccharide binding in family 8 glycosidases: Crystal structures of active-site mutants of the β-1, 4-xylanase pXyl from Pseudoaltermonas haloplanktis TAH3a in complex with substrate and product. Biochemistry 45:4797–4807CrossRefGoogle Scholar
- Pollet A, Lagaert S, Eneyskaya E, Kulminskaya A, Delcour JA, Courtin CM (2010b) Mutagenesis and subsite mapping underpin the importance for substrate specificity of the aglycon subsites of glycoside hydrolase family 11 xylanases. Biochim Biophys Acta Proteins Proteomics 1804:977–985CrossRefGoogle Scholar
- Reilly PJ (1981) Xylanases: structure and function. In: Hollaender A (ed) Trends in the biology of fermentation for fuels and chemicals, basic life sciences. Plenum, New York, pp 111–129Google Scholar
- Takami H, Nakasone K, Takaki Y, Maeno G, Sasaki R, Masui N, Fuji F, Hirama C, Nakamura Y, Ogasawara N, Kuhara S, Horikoshi K (2000) Complete genome sequence of the alkaliphilic bacterium Bacillus halodurans and genomic sequence comparison with Bacillus subtilis. Nucleic Acids Res 28:4317–4331CrossRefGoogle Scholar
- Trogh I, Courtin CM, Goesaert H, Delcour JA, Andersson AAM, Aman P, Fredriksson H, Pyle DL, Sorensen JF (2005) From hull-less barley and wheat to soluble dietary fiber-enriched bread. Cereal Foods World 50:253–260Google Scholar
- Van Craeyveld V, Swennen K, Dornez E, Van de Wiele T, Marzorati M, Verstraete W, Delaedt Y, Onagbesan O, Decuypere E, Buyse J, De Ketelaere B, Broekaert WF, Delcour JA, Courtin CM (2008) Structurally different wheat-derived arabinoxylooligosaccharides have different prebiotic and fermentation properties in rats. J Nutr 138:2348–2355CrossRefGoogle Scholar
- Yoon KH, Yun HN, Jung KH (1998) Molecular cloning of a Bacillus sp. KK-1 xylanase gene and characterization of the gene product. Biochem Mol Biol Int 45:337–347Google Scholar