Sequence Analysis by Cloning of the Structural Gene of Gassericin A, a Hydrophobic Bacteriocin Produced byLactobacillus gasseriLA39

Abstract

Gassericin A, a bacteriocin from Lactobacillus gasseri LA39, was purified to homogeneity from the culturesupernatant mainly by reverse-phase chromatography. The molecular weight of gassericin A was found to be 5,652 by mass analysis, unlike the estimated 3,800 found by SDS-PAGE. However, when the purified preparation was treated with lysylendopeptidase, it migrated as a single band to 5,600 with bacteriocin activity on SDS-PAGE. N- and C-terminal amino acids could not be identified. The internal amino acid could be identified after gassericin A was hydrolyzed with lysylendopeptidase. The DNA of the structural gene of gassericin A was sequenced by cloning of the gene from chromosomal DNA with an oligonucleotide probe. The structural gene of gassericin A was found on the chromosomal DNA as an open reading frame encoding a protein composed of 91 amino acids. The amino acid sequence of mature gassericin A was predicted to be 58 residues from the DNA sequence and results of mass analysis. These results suggested that gassericin A has a closed circular structure with N- and C-terminals linked. Gassericin A is a hydrophobic class II bacteriocin; it was 98% identical with acidocin B produced by Lactobacillus acidophilus M46.