Metabolism of d,l-chloro-phenylalanines by phenylalanine aminotransferase isozymes purified from bushbean shoots

A series of mono-, di- and trichloro-d,l-phenylalanines was tested as substrates for both phenylalanine aminotransferase and phenylalanine decarboxylase partially purified from bushbean (Phaseolus vulgaris L.) seedling extracts by ammonium sulphate fractionation and Sephacryl S-300 gel filtration. While most of the d,l-chlorophenylalanines were transaminated at rates of 35-100% of that observed with d,l-phenylalanine, no chlorophenylalanine decarboxylase activity was observed. A transamination reaction is therefore likely to be the initial step in the conversion of chloro-phenylalanines to their corresponding chloro-phenylacetic acids via a reaction pathway similar to the known route for the metabolism of l-phenylalanine to phenylacetic acid. The highest specific activity of phenylalanine aminotransferase was found in both root and shoot tissues of bushbean at the 10-day stage of seedling growth. Partially purified extracts of these tissues were able to transaminate most of the mono- and dichlorophenylalanines at ca 20-40% of the rate observed with d,l-phenylalanine, while the trichloro-phenylalanines (assayed at lower concentrations due to solubility) were transaminated at rates equal to those observed with d,l-phenylalanine. The 4-chloro derivative was the best substrate tested showing rates of transamination that were 25 % higher than those observed with d,l-phenylalanine. Further purification of shoot fractions by DEAE-Sephacel chromatography resolved the phenylalanine aminotransferase activity into two peaks (enzymes I and II) which on further purification, were found to behave differently during hydrophobic chromatography and PAGE. These results indicated the presence of two isozymic forms of phenylalanine aminotransferase in bushbean shoots and both were found to catalyse transamination of the monochloro-phenylalanines examined in this study.