title: Purification, characterization of GH11 endo-β-1,4-xylanase from thermotolerant Streptomyces sp. SWU10 and overexpression in Pichia pastoris KM71H.
abstract: We have previously described two forms of an endo-β-1,4-xylanase (XynSW2A and XynSW2B) synthesized by thermotolerant Streptomyces sp. SWU10. Here, we describe another xylanolytic enzyme, designated XynSW1. The enzyme was purified to homogeneity from 2 L of culture filtrate. Its apparent molecular mass was 24 kDa. The optimal pH and temperature were pH 5.0 and 40 °C, respectively. The enzyme was stable in a wide pH ranges (pH 1-11), more than 80 % of initial activity remained at pH 2-11 after 16 h of incubation at 4 °C and stable up to 50 °C for 1 h. Xylobiose and xylotriose were the major xylooligosaccharides released from oat spelt xylan by the action of XynSW1, indicating of endo-type xylanase. The complete xynSW1 gene contains 1,011 bp in length and encode a polypeptide of 336 with 41 amino acids of signal peptide. The amino acid sequence analysis revealed that it belongs to glycoside hydrolase family 11 (GH11). The mature xynSW1 gene without signal peptide sequence was overexpressed in Pichia pastoris KM71H. The recombinant XynSW1 protein showed higher molecular mass due to the differences in glycosylation levels at the six N-glycosylation sites in the amino acid sequence and exhibited better physicochemical properties than those of the native enzyme including higher optimal temperature (60 °C), and specific activity, but lower optimal pH (4.0). Because of their stability in a wide pH ranges, both of native and recombinant enzymes of XynSW1, may have potential application in several industries including food, textile, biofuel, and also waste treatment.
authors.affiliation: Department of Biochemistry, Faculty of Medicine, Srinakharinwirot University, 114 Sukhumvit 23, Bangkok 10110, Thailand.
authors.forename: Warin
authors.initials: W
authors.lastname: Deesukon
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Nishimura
authors.forename: Tatsuji
authors.initials: T
authors.lastname: Sakamoto
authors.forename: Wasana
authors.initials: W
authors.lastname: Sukhumsirichart
mesh_terms: D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004187:Disaccharides / Q000737:chemistry; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D015870:Gene Expression; D010843:Pichia / Q000235:genetics; D013302:Streptomyces / Q000201:enzymology*; D013696:Temperature; D014312:Trisaccharides / Q000737:chemistry
chemical_list: D004187:Disaccharides; D014312:Trisaccharides; C515044:xylotriose; D043364:Endo-1,4-beta Xylanases; C004173:xylobiose