oa Journal of Applied Science in Southern Africa - Cloning, sequencing and expression of an endo-?-1,4- glucanase gene of a Bacillus subtilis CHZ1



The endoglucanase gene (celG) from B. subtilis CHZ1 was amplified by PCR, cloned and expressed in Escherichia coli DH5?. The amplified fragment with the structural gene was 2,589 bp and its nucleotide sequence was determined and analysed. The amplified fragment had an open reading frame of 1,524 bp encoding a predicted protein of 56,472 daltons in size with 508 amino acids. celG genes possible ribosomal binding site was identified and it resembled that of B. subtilis ?43 RNA polymerase. The gene contains a signal sequence codifying for a peptide of 38 amino acid residues. A BLASTN search showed 98 percent identities to endo-?-1,4-glucanase that has a predicted protein structure having a cellulases catalytic domain (CD) of 278 amino acids linked to a cellulose binding domain (CBD_3) with 83 residues. The enzyme was identified to belong to family-5A glycosyl hydrolases. There was no drastic reduction in endoglucanase activity during cultivation of the clones as was observed with the wild type strain.


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