title: Novel intracellular GH10 xylanase from Cohnella laeviribosi HY-21: biocatalytic properties and alterations of substrate specificities by site-directed mutagenesis of Trp residues.
abstract: The novel intracellular GH10 xylanase (iXylC) gene (1023-bp) of Cohnella laeviribosi HY-21 encoded a protein consisting of 340 amino acids with a deduced molecular mass of 39,330Da and a calculated pI of 5.81. The primary structure of iXylC was 70% identical to that of Geobacillus sp. GH10 enzyme (GenBank accession number: EDV78425). Xylanolytic activity of the His-tagged iXylC overproduced in Escherichiacoli BL21 was stimulated by 2.2-fold in the presence of 0.5% non-ionic detergents. iXylC produced a mixture of xylooligosaccharides (xylobiose to xylooctaose) from xylotriose and xylotetraose used as the hydrolytic substrate. In addition, it exhibited considerable cleavage activities for p-nitrophenylxylopyranoside (PNP-xylopyranoside) and PNP-cellobioside, indicating that iXylC is a unique GH10 enzyme. The hydrolytic activity (57.8IUmL(-1)) of iXylC toward PNP-xylopyranoside increased to 8.3-fold by W217A and W315A mutations, while mutations of W133A, W295A, and W303A abolished the hydrolytic activity of the enzyme.
authors.affiliation: Industrial Bio-materials Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Republic of Korea.
authors.forename: Do Young
authors.initials: DY
authors.lastname: Kim
authors.forename: Mi Kyoung
authors.initials: MK
authors.lastname: Han
authors.forename: Hyun-Woo
authors.initials: HW
authors.lastname: Oh
authors.forename: Kyung Sook
authors.initials: KS
authors.lastname: Bae
authors.forename: Tae-Sook
authors.initials: TS
authors.lastname: Jeong
authors.forename: Sung Uk
authors.initials: SU
authors.lastname: Kim
authors.forename: Dong-Ha
authors.initials: DH
authors.lastname: Shin
authors.forename: In-Ho
authors.initials: IH
authors.lastname: Kim
authors.forename: Young Ha
authors.initials: YH
authors.lastname: Rhee
authors.forename: Kwang-Hee
authors.initials: KH
authors.lastname: Son
authors.forename: Ho-Yong
authors.initials: HY
authors.lastname: Park
mesh_terms: D002384:Catalysis; D043364:Endo-1,4-beta Xylanases / Q000096:biosynthesis* / Q000235:genetics*; D016297:Mutagenesis, Site-Directed / Q000379:methods*; D056507:Paenibacillus / Q000502:physiology*; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D014364:Tryptophan / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; D014364:Tryptophan; D043364:Endo-1,4-beta Xylanases