Isolation and some properties of an aryl acylamidase from red rice, Oryza sativa L., that metabolizes 3′,4′-dichloropropionanilide1

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Abstract
An aryl acylamidase (aryl-acylamine amidohydrolase, E. C. 3.5.1.a) was isolated from red rice (Oryza sativa L.) seedlings and purified approximately 5-fold with ammonium sulfate precipitation and gel filtration. The enzyme hydrolyzed 3′,4′-dichloro- propionanilide (propanil) and several other mono- and dichloro-substituted propionanilides and 3′,4′-dichloroanilide alkyl analogs. Substrate specificity tests on these analogs indicated the following substrate preference: propanil > 3′-chloropropionanilide > propionanilide > 4′-chloropropionanilide ≥ 3′,5'-dichloropropionanilide ≥ 2'- chioropropionanilide. Tests on a series of 3′,4′-dichloroanilide alkyl analogs of propanil revealed: propanil > 3,4-dichloroacetanilide > 3′,4′-dichlorovaleranilide. The apparent Km for 3′,4′-dichloropropionanilide was determined from a double reciprocal plot of initial velocities at several substrate concentrations. A value of Km=2.5 × 10−5 M was obtained. The temperature optimum, when assayed with propanil as substrate, was 50°C. Although the pH optimum was 8.2, there was relatively high enzyme activity over a wide range of pH values between 7.4–8.7. Several compounds, e.g. p-benzoquinone, pyrocatechol and CuSO4, were found to effectively inhibit the enzyme activity on propanil. These results indicated that the properties of this aryl acylamidase from red rice are very similar to those of enzymes from a commercial rice variety and from other plant species.