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Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max.
Alekseeva, A A; Kargov, I S; Kleimenov, S Yu; Savin, S S; Tishkov, V I.
Afiliación
  • Alekseeva AA; A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia.
  • Kargov IS; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia ; Departament of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory, 1/3, Moscow, 119991, Russia.
  • Kleimenov SY; A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; N.K. Koltsov Institute of Developmental Biology of the Russian Academy of Sciences, Vavilova St., 26, Moscow, 119334, Russ
  • Savin SS; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia ; Departament of Chemistry, M.V. Lomonosov Moscow State University, Leninskie Gory, 1/3, Moscow, 119991, Russia.
  • Tishkov VI; A.N.Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninskiy Prospect, 33/2, Moscow,119071, Russia ; Innovations and High Technologies MSU Ltd, Tsimlyanskaya St., 16, Moscow, 109559, Russia ; Departament of Chemistry, M.V.
Acta Naturae ; 7(3): 55-64, 2015.
Article en En | MEDLINE | ID: mdl-26483960
Recently, we demonstrated that the amino acid substitutions Ala267Met and Ala267Met/Ile272Val (Alekseeva et al., Biochemistry, 2012), Phe290Asp, Phe290Asn and Phe290Ser (Alekseeva et al., Prot. Eng. Des. Select, 2012) in recombinant formate dehydrogenase from soya Glycine max (SoyFDH) lead to a significant (up to 30-100 times) increase in the thermal stability of the enzyme. The substitutions Phe290Asp, Phe290Asn and Phe290Ser were introduced into double mutant SoyFDH Ala267Met/Ile272Val by site-directed mutagenesis. Combinations of three substitutions did not lead to a noticeable change in the catalytic properties of the mutant enzymes. The stability of the resultant triple mutants was studied through thermal inactivation kinetics and differential scanning calorimetry. The thermal stability of the new mutant SoyFDHs was shown to be much higher than that of their precursors. The stability of the best mutant SoyFDH Ala267Met/Ile272Val/Phe290Asp turned out to be comparable to that of the most stable wild-type formate dehydrogenases from other sources. The results obtained with both methods indicate a great synergistic contribution of individual amino acid substitutions to the common stabilization effect.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Acta Naturae Año: 2015 Tipo del documento: Article País de afiliación: Rusia Pais de publicación: Rusia
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Acta Naturae Año: 2015 Tipo del documento: Article País de afiliación: Rusia Pais de publicación: Rusia