Abstract
NAD+-dependent formate dehydrogenases (EC 1.2.1.2, FDH) of methylotrophic bacteria Pseudomonas sp. 101 (PseFDH) and Mycobacterium vaccae N10 (MycFDH) exhibit high homology. They differ in two amino acid residues only among a total of 400, i.e., Ile35 and Glu61 in MycFDH substitute for Thr35 and Lys61 as in PseFDH. However, the rate constant for MycFDH thermal inactivation in the temperature range of 54-65°C is 4-6-times higher than the corresponding rate constant for the enzyme from Pseudomonas sp. 101. To clarify the role of these residues in FDH stability the dependence of the apparent rate constant for enzyme inactivation on phosphate concentration was studied. Kinetic and thermodynamic parameters for thermal inactivation were obtained for both recombinant wild-type and mutant forms, i.e., MycFDH Glu61Gln, Glu61Pro, Glu61Lys and PseFDH Lys61Arg. It has been shown that the lower stability of MycFDH compared to that of PseFDH is caused mainly by electrostatic repulsion between Asp43 and Glu61 residues. Replacement of Lys61 with an Arg residue in the PseFDH molecule does not result in an increase in stability.
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REFERENCES
Rodionov, Yu. V. (1981) Uspekhi Mikrobiol., 16, 104–138.
Popov, V. O., and Lamzin, V. S. (1994) Biochem. J., 301, 625–643.
Vinals, C., Depiereux, E., and Feytmans, E. (1993) Biochem. Biophys. Res. Commun., 192, 182–188.
Egorov, A. M., Avilova, T. V., Dikov, M. M., Popov, V. O., Rodionov, Yu. V., and Berezin, I. V. (1979) Eur. J. Biochem., 99, 569–576.
Asano, Y., Sekigawa, T., Inukai, H., and Nakazawa, A. (1988) J. Bacteriol., 170, 3189–3193.
Iida, M., Kitamura–Kimura, K., Maeda, H., and Mineki, S. (1992) Biosci. Biotech. Biochem., 56, 1966–1970.
Galkin, A., Kulakova, L., Tishkov, V., Esaki, N., and Soda, K. (1995) Appl. Microbiol. Biotechnol., 44, 479–483.
Mitsunaga, T., Tanaka, Y., Yoshida, T., and Watanabe, K. (2000) Japan Patent JP245471A2.
Tishkov, V. I., Galkin, A. G., Marchenko, G. N., Tsygankov, Y. D., and Egorov, A. M. (1993) Biotechnol. Appl. Biochem., 18, 201–207.
Shinoda, T., Satoh, T., Mineki, S., Iida, M., and Taguchi, H. (2002) Biosci. Biotech. Biochem., 66, 271–276.
Barnett, M. J., Fisher, R. F., Jones, T., Komp, C., Abola, A. P., Barloy–Hubler, F., Bowser, L., Capela, D., Galibert, F., Gouzy, J., Gurjal, M., Hong, A., Huizar, L., Hyman, R. W., Kahn, D., Kahn, M. L., Kalman, S., Keating, D. H., Palm, C., Peck, M. C., Surzycki, R., Wells, D. H., Yeh, K.–C., Davis, R. W., Federspiel, N. A., and Long, S. R. (2001) Proc. Natl. Acad. Sci. USA, 98, 9883.
Hollenberg, C. P., and Janowicz, Z. (1989) European Patent EP1987000110417, Bulletin 89/03.
Allen, S. J., and Holbrook, J. J. (1995) Gene, 162, 99–104.
Sakai, Y., Murdanoto, A. P., Konishi, T., Iwamatsu, A., and Kato, N. (1997) J. Bacteriol., 179, 4480–4485.
Slusarczyk, H., Felber, S., Kula, M. R., and Pohl, M. (2000) Eur. J. Biochem., 267, 1280–1289.
Saleeba, J. A., Cobbett, C. S., and Hynes, M. J. (1992) Mol. Gen. Genet., 235, 349–358.
Chow, C. M., and RajBhandary, U. L. (1993) J. Bacteriol., 175, 3703–3709.
Colas des Francs–Small, C., Ambard–Bretteville, F., Small, I. D., and Remy, R. (1993) Plant Physiol., 102, 1171–1177.
Suzuki, K., Itai, R., Suzuki, K., Nakanishi, H., Nishizawa, N. K., Yoshimura, E., and Mori, S. (1998) Plant Physiol., 116, 725–732.
Lamzin, V. S., Dauter, Z., Popov, V. O., Harutyunyan, E. H., and Wilson, K. S. (1994) J. Mol. Biol., 236, 759–785.
Galkin, A. G., Kutsenko, A. S., Bajulina, N. P., Esipova, N. G., Lamzin, V. S., Mezentzev, A. V., Shelukho, D. V., Tikhonova, T. V., Tishkov, V. I., Ustinnikova, T. B., and Popov, V. O. (2002) Biochim. Biophys. Acta, 1594, 136–149.
Tishkov, V. I., Galkin, A. G., Gladyshev, V. N., Karzanov, V. V., and Egorov, A. M. (1992) Biotekhnologiya, No. 5, 52–59.
Rodionova, Yu. V., Avilova, T. V., and Popov, V. O. (1977) Biokhimiya, 43, 2020–2026.
Menendez–Arias, L., and Argos, P. (1989) J. Mol. Biol., 206, 397–406.
Cornish–Bowden, A. (1976) Principles of Enzyme Kinetics, Chap. 1, Butterworth & Co, London–Boston.
Rojkova, A. M., Galkin, A. G., Kulakova, L. B., Serov, A. E., Savitsky, P. A., Fedorchuk, V. V., and Tishkov, V. I. (1999) FEBS Lett., 445, 183–188.
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Fedorchuk, V.V., Galkin, A.G., Yasny, I.E. et al. Effect of Interactions Between Amino Acid Residues 43 and 61 on Thermal Stability of Bacterial Formate Dehydrogenases. Biochemistry (Moscow) 67, 1145–1151 (2002). https://doi.org/10.1023/A:1020915324159
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DOI: https://doi.org/10.1023/A:1020915324159