Cloning and characterization of a xylanase, KRICT PX1 from the strain Paenibacillus sp. HPL-001.
The KRICT PX1 gene (GB: FJ380951) consisting of 996bp encoding a protein of 332 amino acids (38.1kDa) from the recently isolated Paenibacillus sp. strain HPL-001 (KCTC11365BP) has been cloned and expressed in Escherichia coli. The xylanase KRICT PX1 showed high activity on birchwood xylan, and was active over a pH range of 5.0 to 11.0, with two optima at pH 5.5 and 9.5 at 50 degrees C with K(m) value of 5.35 and 3.23, respectively. The xylanase activity was not affected by most salts, such as NaCl, LiCl, KCl, NH(4)Cl, CaCl(2), MgCl(2), MnCl(2), and CsCl(2) at 1mM, but affected by CuSO(4), ZnSO(4), and FeCl(3). One mM of EDTA, 2-mercaptoethanol, and PMSF did not affect the xylanase activity. TLC analysis of the catalyzed products after reaction with birchwood xylan revealed that xylobiose was the major product with smaller amounts of xylotriose and xylose. A similarity analysis of the amino acids in KRICT PX1 resulted 72% identity with xylanase from Geobacillus stearothermophilus (GB: ZP_03040360), 70% identity with intracellular xylanase from an uncultured bacterium (GB: AAP51133), 68% identity with endo-1-4-xylanse from Paenibacillus sp. (GB: ZP_02847150). In addition, the amino acid alignment of KRICT PX1 with glycosyl hydralase (GH) family 10 xylanases revealed a high degree of homology in highly conserved regions including the catalytic sites, and this was confirmed through PROSITE scan. These results imply that KRICT PX1 is a new xylanase gene, and this alkaline xylanase belongs to GH family 10.
Hwang
In Taek
IT
Biorefinery Research Center, Green Chemistry Division, Korea Research Institute of Chemical Technology, Yuseong P.O. Box 107, Daejon 305-600, Republic of Korea.
Lim
Hee Kyung
HK
Song
Ha Young
HY
Cho
Soo Jin
SJ
Chang
Jong-San
JS
Park
No-Joong
NJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D016014:Linear Models; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D056507:Paenibacillus / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012492:Salts / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature
D008670:Metals; D011994:Recombinant Proteins; D012492:Salts; D043364:Endo-1,4-beta Xylanases