1-Aminocyclopropane-1-carboxylate (ACC) deaminases from Methylobacterium radiotolerans and Methylobacterium nodulans with higher specificity for ACC

Authors

  • Dmitry N. Fedorov,

    1. G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms RAS, Pushchino, Moscow region, Russia
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  • Galina A. Ekimova,

    1. G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms RAS, Pushchino, Moscow region, Russia
    2. Pushchino State Institute of Natural Sciences, Pushchino, Moscow region, Russia
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  • Nina V. Doronina,

    1. G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms RAS, Pushchino, Moscow region, Russia
    2. Pushchino State Institute of Natural Sciences, Pushchino, Moscow region, Russia
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  • Yuri A. Trotsenko

    Corresponding author
    1. Pushchino State Institute of Natural Sciences, Pushchino, Moscow region, Russia
    • G.K. Skryabin Institute of Biochemistry and Physiology of Microorganisms RAS, Pushchino, Moscow region, Russia
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Correspondence: Yuri A. Trotsenko, IBPM RAS, Prospekt Nauki, 5, Pushchino, Moscow region, Russia, 142290. Tel.: +7 495 9257448; fax: + 7 495 9563370; e-mail: trotsenko@ibpm.pushchino.ru

Abstract

The 1-aminocyclopropane-1-carboxylate (ACC) deaminases (EC 3.4.99.7), the key enzymes of degradation of the precursor of the phytohormone ethylene, have not been well studied despite their great importance for plant–bacterial interactions. Using blast, the open reading frames encoding ACC deaminases were found in the genomes of epiphytic methylotroph Methylobacterium radiotolerans JCM2831 and nodule-forming endosymbiont Methylobacterium nodulans ORS2060. These genes were named acdS and cloned; recombinant proteins were expressed and purified from Escherichia coli. The enzyme from M. nodulans displayed the highest substrate specificity among all of the characterized ACC deaminases (Km 0.80 ± 0.04 mM), whereas the enzyme from M. radiotolerans had Km 1.8 ± 0.3 mM. The kcat values were 111.8 ± 0.2 and 65.8 ± 2.8 min−1 for the enzymes of M. nodulans and M. radiotolerans, respectively. Both enzymes are homotetramers with a molecular mass of 144 kDa, as was demonstrated by size exclusion chromatography and native PAGE. The purified enzymes displayed the maximum activity at 45–50 °C and pH 8.0. Thus, the priority data have been obtained, extending the knowledge of biochemical properties of bacterial ACC deaminases.

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