Sequence of xynC and properties of XynC, a major component of the Clostridium thermocellum cellulosome

Abstract

The nucleotide sequence of the Clostridium thermocellum F1 xynC gene, which encodes the xylanase XynC, consists of 1,857 bp and encodes a protein of 619 amino acids with a molecular weight of 69,517. XynC contains a typical N-terminal signal peptide of 32 amino acid residues, followed by a 165- amino-acid sequence which is homologous to the thermostabilizing domain. Downstream of this domain was a family 10 catalytic domain of glycosyl hydrolase. The C terminus separated from the catalytic domain by a short linker sequence contains a dockerin domain responsible for cellulosome assembly. The N-terminal amino acid sequence of XynC-II, the enzyme purified from a recombinant Escherichia coli strain, was in agreement with that deduced from the nucleotide sequence although XynC-II suffered from proteolytic truncation by a host protease(s) at the C-terminal region. Immunological and N-terminal amino acid sequence analyses disclosed that the full-length XynC is one of the major components of the C. thermocellum cellulosome. XynC-II was highly active toward xylan and slightly active toward p-nitrophenyl-β-D-xylopyranoside, p-nitrophenyl-β-D-cellobioside, p- nitrophenyl-β-D-glucopyranoside, and carboxymethyl cellulose. The K(m) and V(max) values for xylan were 3.9 mg/ml and 611 μmol/min/mg of protein, respectively. This enzyme was optimally active at 80°C and was stable up to 70°C at neutral pHs and over the pH range of 4 tn 11 at 25°C.