Presence of Active and Inactive Molecules of a Cell Wall-Associated Proteinase in Lactobacillus helveticus CP790

Author:

Yamamoto N,Akino A,Takano T,Shishido K

Abstract

Monoclonal antibodies against a cell wall-associated 45-kDa proteinase from Lactobacillus helveticus CP790 were prepared and used for an immunoblotting analysis of the cell wall extract of CP790. They were found to react with an unidentified 46-kDa protein as well as the 45-kDa proteinase. The 46-kDa protein was copurified with the 45-kDa proteinase by affinity column chromatography using antibody-fixed Sepharose and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then extracted from the gels. An elution profile of the cyanogen bromide digest of the purified 46-kDa protein obtained by reversed-phase high-performance liquid chromatography was identical to that of the 45-kDa proteinase except for one peak. An analysis of the N-terminal 21-amino-acid sequence revealed that the 46-kDa protein possesses an extra 7 amino acids at the N terminus of the 45-kDa proteinase. The 46-kDa protein was produced at constant levels during fermentation in a skim milk medium, while the 45-kDa protein was mainly observed in the middle of the exponential phase of growth and was produced in proportion to the proteinase activity. Moreover, only the 46-kDa protein was detected in the crude extract of L. helveticus CP791, a variant strain of CP790 defective in proteinase activity. These data strongly suggest that the 46-kDa protein is a precursor, inactive form of the 45-kDa proteinase.

Publisher

American Society for Microbiology

Subject

Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology

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