Molecular Cloning, Isolation, and Properties of Chaperone Skp from Yersinia pseudotuberculosis

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E. V. Sidorin^*, N. M. Tishchenko, V. A. Khomenko, M. P. Isaeva, P. S. Dmitrenok, N. Yu. Kim, G. N. Likhatskaya, and T. F. Solov’eva

Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch of the Russian Academy of Sciences, pr. 100 Let Vladivostoku 159, 690022 Vladivostok, Russia; fax: (423) 231-4050; E-mail: sev1972@mail.ru

^* To whom correspondence should be addressed.

Received May 29, 2012; Revision received June 25, 2012

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The skp gene of Yersinia pseudotuberculosis was expressed without its signal sequence in Escherichia coli BL21(DE3) cells. The recombinant protein Skp accumulated in soluble form in the cytoplasm of the producer strain. The protein was isolated and characterized: the molecular weight, isoelectric point, N-terminal amino acid sequence (20 amino acid residues), and the content of the secondary structure elements were determined. Using cross-linking stabilization and high-mass MALDI-TOF mass spectrometric analysis, it was found that rSkp forms a stable homotrimer in solution and interacts with human IgG. Three-dimensional models of the Skp trimer and its complexes with Fc- and Fab-fragments of human IgG1 were constructed by computer modeling.

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KEY WORDS: chaperone Skp, Yersinia pseudotuberculosis, immunoglobulin G, cross-linking stabilization, MALDI-TOF mass spectrometry, computer modeling, protein–protein interactions

DOI: 10.1134/S0006297912110119

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