Purification and characterization of a lecithin-dependent haemolysin from Escherichia coli transformed by a Vibrio parahaemolyticus gene.
Lecithin-dependent haemolysin (LDH) of Vibrio parahaemolyticus was purified from Escherichia coli C600 transformed with a plasmid (pHL591) ligated with a 1.5 kb DNA fragment of V. parahaemolyticus. The final preparation comprised two LDH proteins with different molecular masses which were immunologically cross-reactive and had the same enzymic activity. The LDH was a phospholipase hydrolysing both fatty acid esters of phospholipid, i.e. it hydrolysed phosphatidylcholine (PC) to lysophosphatidylcholine (LPC) and then LPC to glycerophosphorylcholine (GPC). From this point of view, LDH should be classified as a phospholipase B. Phospholipase B, however, does not usually show haemolytic activity, because the intermediate (LPC), which is the actual haemolytic agent, is immediately hydrolysed to the final product (GPC). On the other hand, LPC formed by LDH action was comparatively stable, because the rates of the two reactions catalysed by LDH, PC to LPC and LPC to GPC, are almost the same. This is the reason that LDH shows haemolytic activity. Therefore, LDH of V. parahaemolyticus is an atypical phospholipase to be designated as phospholipase A2/lysophospholipase.
Shinoda
S
S
Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Matsuoka
H
H
Tsuchie
T
T
Miyoshi
S
S
Yamamoto
S
S
Taniguchi
H
H
Mizuguchi
Y
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D003429:Cross Reactions; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D008244:Lysophosphatidylcholines / Q000378:metabolism; D008969:Molecular Sequence Data; D010713:Phosphatidylcholines / Q000378:metabolism*; D010740:Phospholipases / Q000378:metabolism; D013379:Substrate Specificity; D014169:Transformation, Bacterial; D014736:Vibrio parahaemolyticus / Q000235:genetics*
D004269:DNA, Bacterial; D006460:Hemolysin Proteins; D008244:Lysophosphatidylcholines; D010713:Phosphatidylcholines; D010740:Phospholipases