Enzymatic Properties of Thiol-Dependent Serine Proteinase of Bacillus intermedius 3-19

E. L. Itskovich,¹ N. P. Balaban,¹ A. M. Mardanova,¹ E. V. Shakirov,¹ M. R. Sharipova,¹ I. B. Leshchinskaya,¹ A. L. Ksenofontov,² and G. N. Rudenskaya^2,3

¹Department of Microbiology, School of Biology, Kazan State University, ul. Lenina 18, Kazan, 420008 Russia; fax: (8-432) 38-09-94; E-mail: Nelly.Balaban@ksu.ru

²Department of Chemistry of Natural Compounds, School of Chemistry, Lomonosov Moscow State University, Moscow, 119899 Russia; fax: (7-095) 939-31-81; E-mail: rudenskaya@biorg.chem.msu.su

³To whom correspondence should be addressed.

Submitted May 30, 1996; revision submitted October 11, 1996.

Effects of a thiol-dependent serine proteinase of Bacillus intermedius on peptide substrates and insulin B-chain were studied. The enzyme preferably splits peptide bonds formed by carboxyl groups of hydrophobic amino acids. Ca²⁺ increases the thermal stability of the proteinase significantly. The kinetic characteristics of hydrolysis of Z-Ala-Ala-Leu-pNA by this enzyme was determined as K_m = 1.25 mM and k_cat = 0.15 sec^-1. The enzyme has high stability to DMFA and isopropanol, and is able to catalyze peptide bond synthesis.

KEY WORDS: thiol-dependent serine proteinase, substrate specificity, enzymatic synthesis of substrates.

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