Molecular cloning, sequence analysis and expression of the catalase-peroxidase gene from Halobacterium salinarium.

摘要

The complete catalase-peroxidase, gene was cloned from chromosomal DNA from the Archaea, Halobacterium salinarium. The nucleotide sequence of a 3.5 kb fragment, containing the catalase-peroxidase gene and its flanking regions was determined. A 2,160 bp open reading frame, coding for a catalase-peroxidase of 720 amino, acid residues with a calculated molecular weight of 80 kDa, was obtained. The amino acid sequence of H. salinarium catalase-peroxidase showed a high degree of similarity to other bifunctional catalase-peroxidases. A transcriptional start site was identified which was 183 bp upstream of the translational start codon. Southern blot analysis indicated that catalase-peroxidase was a single copy gene. The expression of catalase-peroxidase in H. salinarium was not induced by H2O2, redox inhibitors, heavy metals, ions, light or temperature. The catalase-peroxidase gene has been expressed in Escherichia coli, yeast and the halophilic Archaea, H. salinarium. The enzyme activity increased 2-fold when the catalase-peroxidase gene with its own upstream promoters was transformed into H. salinarium. The enzyme activity increased 3--fold when the catalase-peroxidase gene under control of a halobacterial tRNALys(HTP) promoter was transformed into H. salinarium .