P54: Novel isonitrile hydratase involved in isonitrile metabolism in Arthrobacter sp

We previously discovered N-substituted formamide deformylase (NfdA) in Arthrobacter pascens F164, which degrades N-substituted formamide [PNAS 101, 13726 (2004)]. We here found an enzyme involved in the first step of isonitrile metabolism: isonitrile hydratase, which hydrates isonitrile to the corresponding N-substituted formamide. We at first investigated the optimum culture conditions for the enzyme production. The highest enzyme activity was obtained when the strain was cultured in a nutrient medium with N-benzylformamide. This enzyme (named InhB) was purified, characterized and compared with Pseudomonas putida N19-2 isonitrile hydratase (InhA). InhB had a molecular mass of about 530 kDa and consisted of twelve identical subunits. InhB showed different preference to substrate (isonitrile) from InhA. Comparison of these properties between InhB and InhA and Western blot analysis demonstrated that both enzymes are biochemically and immunologically different each other. A. pascens F164 grew and exhibited the isonitrile hydratase and N-substituted formamide deformylase activities when cultured in a medium containing isonitrile as the sole carbon and nitrogen sources. However, both enzyme activities were not induced when cultured in a medium containing glycerol and (NH₄)₂SO₄ as the sole carbon and nitrogen sources. These findings demonstrated that the physiological role of InhB is assimilation of isonitrile in contrast with detoxification as the biological function of InhA.