Molecular analysis of algicidal activity of extracellular serine protease produced by Pseudoalteromonas sp. strain A28

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The marine bacterium Pseudoalteromonas sp. strain A28 produces an extracellular serine protease, designated protease I, to kill the red-tide alga Skeletonema costatum. The espI gene encoding protease I was cloned and completely sequenced. DNA sequence analysis revealed that protease I undergoes removals of 148-amino-acid and 4.6-kDa polypeptides from its amino and carboxy termini, respectively, to form the mature protein. The mature protease I contains two repeated sequences of 55 amino acid residues in the carboxy terminal region. A series of truncated genes of espI, encoding C-terminal truncation mutants of protease I, were generated. Transformants containing die truncated genes showed poor growth (Escherichia coli) or no growth (Pseudoalteromonas sp.). The Bacillus subtilis transformants produced and excreted the truncation mutants. A skim-milk-polyacrylamide gel electrophoresis protease assay showed that these truncation mutants had a proteolytic activity. Paper disk assays revealed that these deletion mutant proteins showed algicidal activity in the assay conditions