title: Molecular and biochemical characterization of a novel intracellular low-temperature-active xylanase.
abstract: A 990 bp full-length gene (xynAHJ2) encoding a 329- residue polypeptide (XynAHJ2) with a calculated mass of 38.4 kDa was cloned from Bacillus sp. HJ2 harbored in a saline soil. XynAHJ2 showed no signal peptide, distinct amino acid stretches of glycoside hydrolase (GH) family 10 intracellular endoxylanases, and the highest amino acid sequence identity of 65.3% with the identified GH 10 intracellular mesophilic endoxylanase iM-KRICT PX1-Ps from Paenibacillus sp. HPL-001 (ACJ06666). The recombinant enzyme (rXynAHJ2) was expressed in Escherichia coli and displayed the typical characteristics of low-temperatureactive enzyme (exhibiting optimum activity at 35 degrees C, 62% at 20 degrees C, and 38% at 10 degrees C; thermolability at > or =45 degrees C). Compared with the reported GH 10 low-temperature-active endoxylanases, which are all extracellular, rXynAHJ2 showed low amino acid sequence identities (<45%), low homology (different phylogenetic cluster), and difference of structure (decreased amount of total accessible surface area and exposed nonpolar accessible surface area). Compared with the reported GH 10 intracellular endoxylanases, which are all mesophilic and thermophilic, rXynAHJ2 has decreased numbers of arginine residues and salt bridges, and showed resistance to Ni2+, Ca2+, or EDTA at 10 mM final concentration. The above mechanism of structural adaptation for low-temperature activity of intracellular endoxylanase rXynAHJ2 is different from that of GH 10 extracellular low-temperature-active endoxylanases. This is the first report of the molecular and biochemical characterizations of a novel intracellular low-temperatureactive xylanase.
authors.affiliation: Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education, Yunnan Normal University, Kunming 650500, People's Republic of China.
authors.forename: Junpei
authors.initials: J
authors.lastname: Zhou
authors.forename: Yanyan
authors.initials: Y
authors.lastname: Dong
authors.forename: Xianghua
authors.initials: X
authors.lastname: Tang
authors.forename: Junjun
authors.initials: J
authors.lastname: Li
authors.forename: Bo
authors.initials: B
authors.lastname: Xu
authors.forename: Qian
authors.initials: Q
authors.lastname: Wu
authors.forename: Yajie
authors.initials: Y
authors.lastname: Gao
authors.forename: Lu
authors.initials: L
authors.lastname: Pan
authors.forename: Zunxi
authors.initials: Z
authors.lastname: Huang
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000737:chemistry / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D003080:Cold Temperature; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D012988:Soil Microbiology
chemical_list: D001426:Bacterial Proteins; D043364:Endo-1,4-beta Xylanases