Cloning, sequencing, and expression of the gene encoding the Clostridium stercorarium xylanase C in Escherichia coli.
The nucleotide sequence of the Clostridium stercorarium F-9 xynC gene, encoding a xylanase XynC, consists of 3,093 bp and encodes a 1,031-amino acids with a molecular weight of 115,322. XynC is a multidomain enzyme composed of an N-terminal signal peptide and six domains in the following order: two thermostabilizing domains, a family 10 xylanase domain, a family IX cellulose-binding domain, and two S-layer homologous domains. Immunological analysis indicated the presence of XynC in the culture supernatant of C. stercorarium F-9 and in the cells, most likely on the cell surface. XynC purified from a recombinant E. coli was highly active toward xylan and slightly active toward p-nitrophenyl-beta-D-xylopyranoside, p-nitrophenyl-beta-D-cellobioside, p-nitrophenyl-beta-D-glucopyranoside, and carboxymethylcellulose. XynC hydrolyzed xylan and xylooligosaccharides larger than xylotriose to produce xylose and xylobiose. This enzyme was optimally active at 85 degrees C and was stable up to 75 degrees C at pH 5.0 and over the pH range of 4 to 7 at 25 degrees C.
Ali
M K
MK
Department of Bioscience, Faculty of Bioresources, Mie University, Tsu, Japan.
Fukumura
M
M
Sakano
K
K
Karita
S
S
Kimura
T
T
Sakka
K
K
Ohmiya
K
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D043364:Endo-1,4-beta Xylanases; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D007120:Immunochemistry; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D017931:DNA Primers; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases