Analysis of phospholipase D gene from Streptoverticillium reticulum and the effect of biochemical properties of substrates on phospholipase D activity

摘要

The phospholipase D (PLD) gene encoding the mature PLD enzyme was cloned from the genomic DNA library of Streptoverticillium reticulum and its gene sequence was analyzed.The Stv.reticulum PLD gene sequence showed a high degree of similarity to that of Strepto-myces spp.(66%),Streptomyces antibioticus (67%),Streptoverticillium cinnamoneum (70%),and Streptomyces somaliensis (71%).A highly conserved phospholipase superfamily-specific consensus sequence,HxKxxxxD (HKD),was also found in both N- and C-terminal regions of the peptides from Streptoverticillium reticulum.The effects of biochemical properties of phosphatidylcholines (PC) on Streptoverticillium reticulum PLD activity were studied.The biochemical properties of PC studied include the chain length,cis and trans conformation of PC,carboxyl and carbonyl bonds at position 1 and 2 of PC,hydrophobic chain of PC,and saturated and unsaturated acyl and alkyl groups.Also studied are the micelle sizes of PC and mixed TX-100-PC,the diacyl chain length,and the properties of various functional groups in the presence and absence of TX-100.The PLD activity was significantly affected by the biochemical properties of PC substrates used and the micelle size.A prospective reaction mechanism of this enzyme has been also suggested.The PLD activity was significantly affected by the biochemical properties of PC substrates used and the micelle size.A prospective reaction mechanism of this enzyme has been also suggested.The studies on the Stv.reticulum PLD gene analysis and the effect of PC substrates in PLD activity are important not only for better understanding of PLD function in cells but also for production of phospholipid derivatives used in pharmaceutical,food,and cosmetics industries.