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D. A. Gudkov, I. V. Lyagin, V. V. Verkhusha, Ye. N. Efremenko
New chimeric proteins
possessing organophosphoroushydrolase
activity and deGFP4 fluorescence
Abstract
New
genetic constructs encoding synthesis of chimeric
proteins possessing organophosphoroushydrolase activity and fluorescence of pH-sensitive
analogue of green fluorescent protein were developed. To obtain maximal yield
of new chimeric proteins in soluble form in E.
coli cells, the following conditions were revealed to be used: 0.1 mM
of inductor of protein biosynthesis, cultivation of cells for 10 h after
biosynthesis induction. Obtained level of biosynthesis of one of the chimeric proteins in soluble active form was 2-25-fold
higher than the yields of soluble forms of analogue chimeric
proteins described in literature. It was found that organophosphoroushydrolase as a part of the chimeric
proteins possess features (pH-optimum, thermostability,
substrate specificity and catalytic efficiency of action) differing from known
characteristics of native enzyme. The fluorescence of green fluorescent protein
being part of chimeric proteins depends on pH of
medium by same manner known for fluorescence of the individual protein. The
interrelation between fluorescence and OPH-activity possessing by obtained chimeric proteins was demonstrated in the reactions of
hydrolysis of organophosphorous compounds.
Copyright (C) Chemistry Dept., Moscow State University, 2002
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