Coexpression of All Constituents of the Cholesterol Hydroxylase/Lyase System in Escherichia coli Cells

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T. V. Shashkova^*, V. N. Luzikov, and L. A. Novikova

Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia; fax: (495) 939-3181; E-mail: tshashkova@yandex.ru

^* To whom correspondence should be addressed.

Received February 6, 2006; Revision received March 29, 2006

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Using the pTrc99A/P450scc vector, a plasmid was constructed in which cDNAs for cytochrome P450scc, adrenodoxin reductase, and adrenodoxin are situated in a single expression cassette. This plasmid was shown to direct the synthesis of all the above proteins in Escherichia coli. Their localization in the E. coli cells and stoichiometry were determined. Cell homogenates exhibited cholesterol hydroxylase/lyase activity, due to catalytically active forms of all three proteins. Thus, the full set of constituents of the mammalian cholesterol hydroxylase/lyase system was shown to be synthesized in bacterial cells for the first time.

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KEY WORDS: cytochrome P450scc, adrenodoxin, adrenodoxin reductase, coexpression, Escherichia coli

DOI: 10.1134/S0006297906070145

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