I87E Mutation Prevents Barstar Dimerization |
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| Authors: | Korchuganov D S Schulga A A Ermolyuk Ya S Mitkevich V A Reibarkh M Ya Nolde S B Makarov A A Arseniev A S Kirpichnikov M P |
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| Institution: | (1) Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, Moscow, 117997, Russia;(2) Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, ul. Vavilova 32, Moscow, 119991, Russia;(3) The Center for Medical Studies at Moscow, University of Oslo, ul. Vavilova 34/5, Moscow, 119991, Russia |
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| Abstract: | The C40,82A;I87E mutant of barstar, an intracellular inhibitor of the ribonuclease barnase from Bacillus amyloliquefaciens, was obtained, and its physicochemical properties were studied. It was produced as a fusion protein with thioredoxin and then cleaved from this by EKmax enterokinase. The mutant was shown by NMR to retain the spatial structure of the wild-type protein but, in contrast to barstar, does not form the homodimers characteristic of barstar in aqueous solution. The mutant protein binds barnase with the dissociation constant (6.6 ± 1.1) × 10–11 M and exhibits other physicochemical properties similar to those of the wild-type barstar. This allows the use of C40,82A;I87E mutant instead of wild-type barstar in the investigations where the protein dimerization is undesirable. |
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| Keywords: | barnase barstar protein dimerization |
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