A FACILE TRANSPHOSPHATIDYLATION REACTION USING A CULTURE SUPERNATANT OF ACTINOMYCETES DIRECTLY AS A PHOSPHOLIPASE-D CATALYST WITH A CHELATING AGENT

An attempt was made to use the phospholipase D (PLD)containing culture supernatants of actinomycetes directly as catalysts for the transphos phatidylation reaction of phosphatidylcholine (PC) to phosphatidyletha nolamine (PE) in a biphasic system. Of the five actinomycetes (three S treptomyces sp. and two Streptoverticillium sp.) examined, three (St. mediocidicus, Stv. cinnamoneum, and Stv. hachijoense) exhibited good P LD production performance, but the selectivity (ratio of transphosphat idylation to hydrolysis) of the PLDs in the culture supernatant of all three actinomycetes were significantly low. However, the addition of EDTA to the reaction mixture as a chelating agent remarkably improved the selectivity of the PLDs, which approached 100% in all the culture supernatants. Commercially available PLDs were also investigated and c lassified into two types. The PLDs of one type had high selectivity an d no metal was required for the enzyme activity, while those of the ot her type showed low selectivity and a metal was necessary for the enzy me to be activated. From this finding, it was considered that the cult ure supernatants used in this study contained several PLDs of both typ es. When the chelating agent was added to the reaction mixture, the hy drolysis due tb PLDs with low selectivity was suppressed by removal of the essential metal, resulting in an increase in the overall selectiv ity of the PLDs in the culture supernatant. Repeated batch transphosph atidylation reactions were performed 20 times, reusing the PLDs in the aqueous phase by centrifugation; the reaction rate gradually decrease d to 60% of that of batch 1 by batch 20. Th is suggests that the trans phosphatidylation reaction using a culture supernatant has potential f or industrial application. (C) 1994 John Wiley and Sons, Inc.