Journal of Basic Microbiology 2006-01-01

Purification and characterization of a halotolerant intracellular protease from Bacillus subtilis strain FP-133.

Endang Setyorini, Young-Ju Kim, Shinji Takenaka, Shuichiro Murakami, Kenji Aoki

Index: J. Basic Microbiol. 46 , 294-304, (2006)

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Abstract

A halotolerant strain FP-133, able to grow at concentrations of 0-12.5% (w/v) NaCl, was isolated from a fish paste and identified as Bacillus subtilis . B. subtilis strain FP-133 produced an intracellular protease which showed catalytic activity under saline conditions. The enzyme was purified to homogeneity 143-fold with a yield of 0.9%. The purified enzyme showed an optimum activity at a concentration of 5% (w/v) NaCl. After storage in 7.5% (w/v) NaCl at 4 degrees C for 24 h, the enzyme kept 100% of its activity. The molecular mass of the protease was determined to be 59 kDa by gel filtration; the protein consisted of four subunits each with a molecular mass of 14 kDa. The enzyme showed aminopeptidase activity. It acted on L-leucyl-p-nitroanilide, L-leucyl-beta-naphthylamide, and oligopeptides containing glycine, L-histidine, or L-leucine. The K(m ) and V (max) values for L-leucyl-p-nitroanilide were 18 microm and 2.2 mm/h mg, respectively. The enzyme was activated by Fe(2+), Fe(3+), and Ni(2+) in synergism with Mg(2+).((c) 2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).