Cloning of a Bacillus subtilis AMX-4 xylanase gene and characterization of the gene product.
A gene encoding the xylanase of Bacillus subtilis AMX-4 isolated from soil was cloned into Escherichia coli, and the gene product was purified from the cell-free extract of the recombinant strain. The gene, designated xylA, consisted of 639 nucleotides encoding a polypeptide of 213 residues. The deduced amino acid sequence was highly homologous to those of xylanase belonging to glycosyl hydrolase family 11. The molecular mass of the purified xylanase was 23 kDa as estimated by SDS-PAGE. The enzyme had a pH optimum at 6.0-7.0 and a temperature optimum at 50-55 degrees C. Xylanase activity was significantly inhibited by 5 mM Cu2+ and 5 mM Mn2+, and noticeably enhanced by 5 mM Fe2+. The enzyme was active on xylans including arabinoxylan, birchwood xylan, and oat spelt xylan, but it did not exhibit activity toward carboxymethylcellulose or p-nitrophenyl-beta-xylopyranoside. The predominant products resulting from xylan and xylooligosaccharide hydrolysis were xylobiose and xylotriose. The enzyme could hydrolyze xylooligosaccharides larger than xylotriose.
Yoon
Ki-Hong
KH
Department of Food Science and Biotechnology, Woosong University, 17-2, Jayang-dong, Dong-gu, Daejeon 300-718, Korea. ykh@wsu.ac.kr
D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004187:Disaccharides / Q000096:biosynthesis; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D019216:Metals, Heavy / Q000737:chemistry; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012988:Soil Microbiology; D013379:Substrate Specificity; D013696:Temperature; D014312:Trisaccharides / Q000096:biosynthesis; D014990:Xylans / Q000378:metabolism
D004187:Disaccharides; D019216:Metals, Heavy; D011994:Recombinant Proteins; D014312:Trisaccharides; D014990:Xylans; C515044:xylotriose; D043364:Endo-1,4-beta Xylanases; C004173:xylobiose