Title
Purification and pressure dependence of alanine racemase from the psychro- piezophilic bacterium shewanella violacea DSS 12
Conference Dates
September 15-19, 2019
Abstract
Shewanella violacea DSS12 (S. violacea) is a psychrophilic and piezophilic bacterium, isolated from mud of the Ryukyu Trench in Japan. The bacterium displays optimal growth at 8°C and 30 MPa. Alanine racemase is an enzyme which catalyses the interconversion of l-alanine and d-alanine, and is responsible for the synthesis of d-alanine contained in the peptidoglycan of bacterial cell wall. In this study, we purified alanine racemase from S. violacea and investigated the enzymological characteristics of alanine racemase.
The bacterium was aerobically cultured using marine broth 2216 in a 5-liter medium bottle at 4°C for 3 days. The bacterial cells were lysed by applying of 100 MPa pressure using a French press, and the lysate was centrifuged. The supernatant obtained was ultracentrifuged at 141,000 g, and the supernatant obtained was applied to ammonium sulfate fractionation. The active fraction was dissolved and passed through a butyl-Toyopearl, phenyl-Sepharose, and shodex KW-200 columns to obtain a partially purified enzyme. Consequently, the enzyme was purified 540-fold and showed a specific activity of 2.68 μmol/min/mg. Alanine racemase exhibited high activity against l-Ala and l-Ser as substrates. The optimal pH and temperature of alanine racemase were 9.0 and 25°C, respectively.
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Recommended Citation
Minoru Tanigawa, Keisuke Itabashi, Katsushi Nishimura, Mayumi Tokuhisa, Miya Koike, and Chiaki Kato, "Purification and pressure dependence of alanine racemase from the psychro- piezophilic bacterium shewanella violacea DSS 12" in "Enzyme Engineering XXV", Huimin Zhao, University of Illinois at Urbana-Champaign, USA John Wong, Pfizer, USA Eds, ECI Symposium Series, (2019). https://dc.engconfintl.org/enzyme_xxv/43