Cloning and sequence analysis of the gene for glucodextranase from Arthrobacter globiformis T-3044 and expression in Escherichia coli cells

The gld gene for glucodextranase from Arthrobacter globiformis T-3044 was c loned by using a combination of gene walking and probe methods and expresse d on the recombinant plasmid pGD8, which was constructed with pUC118, in Es cherichia coil cells. The enzyme gene consisted of a unique open reading fr ame of 3,153 bp. The comparison of the DNA sequence data with the N-termina l and 6 internal amino acid sequences of the purified enzyme secreted from A. globiformis T-3044 suggested the enzyme was translated from mRNA as a se cretory precursor with a signal peptide of 28 amino acids residues. The ded uced amino acids sequence of the mature enzyme contained 1,023 residues, re sulting in a polypeptide with a molecular mass of 107,475 daltons. The dedu ced sequence showed about 38% identity to that of the glucoamylase from Clo stridium sp. G0005. The glucodextranase activity of transformant harboring pGD8 was about 40 mU/ml at 30 degrees C for a 16-h culture. Although the GD ase that was produced from the transformant was shorter than authentic GDas e by 2 amino acid residues at the N-terminal end side, its enzymatic proper ties were almost same as the authentic one. Two kinds of genes, dex1 and dex2, for endo-dextranases from A. globiformis T-3044 were also cloned into Escherichia coli cells. The N-terminal of the purified endo-dextranase from A. globiformis T-3044 agreed with the deduce d amino acid sequence, after the 33rd alanine residue, of only the dex1 gen e for endo-dextranase. This result suggests that the endo-dextranase is tra nslated from mRNA as a secretory precursor with a signal peptide of 32 amin o acids residues. The deduced sequence of endo-dextranase 1 and endo-dextra nase 2 showed about 93% and 65% identity with that of known endo-dextranase from Arthrobacter sp. CB-8, respectively.