Isolation of Milk-Clotting Enzyme from Transgenic Sheep Milk and Its Comparison with Calf Chymosin

M. N. Mezina¹, G. I. Lavrenova², M. I. Prokof'ev¹, V. V. Starovoitova^2*, V. I. Ermolaev¹, V. Ya. Chernyh¹, G. N. Balandina², and S. S. Demidovich³

¹Research and Production Animal Biotechnology Center, ul. Administrativnaya 2, Poselok Gorki Leninskie, Moscow Region, 142712 Russia; fax: (095) 548-9232

²Department of Natural Compounds Chemistry, School of Chemistry, Lomonosov Moscow State University, Moscow, 119899 Russia; fax: (095) 932-8846; E-mail: violst@genebee.msu.su

³Moscow Chymosin Factory, Moscow, Russia

^* To whom correspondence should be addressed.

Received June 5, 2000; Revision received June 22, 2000

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Technology for preparation of chymosin from milk of transgenic sheep has been elaborated. Purification of the preparation by ion-exchange chromatography on aminosilochrom and biospecific chromatography on bacitracin-Sepharose yielded homogeneous active enzyme. Hydrolysis of protein substrates (hemoglobin, BSA, and sodium caseinate) by the transgenic sheep chymosin and stability of the enzyme at various values of pH were studied. Judging by the amino acid composition, the N-terminal sequence involving six amino acid residues, molecular mass, stability at various pH values, and the catalytic activity against the protein substrates, the transgenic sheep chymosin is identical to calf chymosin.

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KEY WORDS: transgenic sheep chymosin, calf chymosin

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