NUCLEOTIDE-SEQUENCES OF THE CELB GENE ENCODING ENDO-1,4-BETA-GLUCANASE-2, ORF1 AND ORF2 FORMING A PUTATIVE CELLULASE GENE-CLUSTER OF CLOSTRIDIUM-JOSUI

The nucleotide sequence of the celB gene coding for endo-1,4-beta-glucanase-2 (EG-2) of Clostridium josui was determined. The structural gene consists of an open reading frame of 1,380 bp encoding a 460-amino acid polypeptide. The deduced amino acid sequence of EG-2 shows high similarities with the Clostridium cellulolyticum CelCCC (92.3%) and the Clostridium thermocellum endoglucanase CelA (61.6%) in the catalytic domain, but no homology with EG-1 of C. josui. In the COOH-terminal region of EG-2, there exists a reiterated segment similar to the segments present in many clostridial cellulases and hemicellulases. Upstream and downstream of the celB gene, two incomplete open reading frames (ORF1 and ORF2) were found to encode proteins homologous to the proteins which consist of the C. cellulolyticum cellulase gene cluster. ORF1 containing a reiterated segment showed high similarities with the proteins deduced from each ORF1 from C. cellulolyticum and Caldocellum saccharolyticum. Downstream of the celB gene, ORF2 was found to encode a protein homologous to CelCCG of C. cellulolyticum and CelF of C. thermocellum. These results indicate that the celB gene, ORF1 and ORF2 formed a cellulase gene cluster strictly homologous to that of C. cellulolyticum, which is proposed to be part of a single operon.