Molecular cloning and sequence analysis of a gene encoding an extracellular proteinase from lactobacillus helveticus CP790

Abstract

A 2.6-kilobase HaeIII DNA fragment corresponding to an extracellular proteinase gene (prtY) was cloned from chromosomal DNA of Lactobacillus helveticus CP790 in Escherichia coli using a pKK223-3 vector. The transformant expressed a 48-kDa protein that reacts with monoclonal antibodies specific to the proteinase and seemed to be a pre-proproteinase, but had no proteolytic activity. About 1.6 kilobases of the 2.6-kilobase DNA fragment, which contained the complete gene for the proteinase was sequenced. Sequence analysis found an open reading frame with a capacity to encode a protein of 449 amino acids. The coding region contained a Gram-positive-type signal peptide of 30 amino acids. The N-terminal sequences of the proproteinase and the mature proteinase have been observed in the polypeptide at position +31 and +38. The putative amino acid sequence showed a significant similarity to a surface layer protein of L. helveticus and Lactobacillus acidophilus in the amino terminal signal sequence and carboxyl terminus. © 2000, Taylor & Francis Group, LLC. All rights reserved.