Melittin-mediated release of [3H]-oleic acid from E. coli cells is dependent upon heat- and trypsin-sensitive factor(s) in human serum.
Synthetic melittin mediated the release of [3H]-oleic acid ([3H]-OA) or its acylated lipids from [3H]-OA-labeled E. coli cells exposed to human serum. This phenomenon was not observed in the absence of serum and was calcium independent. The addition of serum was not required for melittin-mediated lysis of erythrocytes, although lysis was greater in the presence of serum than in its absence (P<0.001). Trypsin treatment of human serum reduced the melittin-mediated release of [3H]-OA/acylated lipids, and this effect was more pronounced upon boiling the serum (P<0.01). A kinetic study showed that maximum release of [3H]-OA/acylated lipids occurred within 3-6 min. Thin layer chromatography (TLC) analysis showed the lipids to be phosphatidyl ethanolamine (PE), phosphatidylethanol (PEt) and phosphatidic acid (PA). There was no detectable level of oleic acid (OA), diacylglycerol (DAG), phosphatidyl choline (PC) or phosphatidyl serine (PS). These findings suggested that a trypsin and heat-sensitive enzyme/factor present in the serum had a role in melittin-mediated action. These findings further showed that melittin activated phospholipase D (PLD), without affecting phospholipase A(2) (PLA(2)) or phospholipase C (PLC) activity.
Saini
S S
SS
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX 77555-1070, USA. sssaini@utmb.edu
Chopra
A K
AK
Peterson
J W
JW
D002855:Chromatography, Thin Layer; D004912:Erythrocytes / Q000187:drug effects / Q000201:enzymology; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism*; D006461:Hemolysis / Q000187:drug effects; D006358:Hot Temperature; D006801:Humans; D007202:Indicators and Reagents; D008055:Lipids / Q000737:chemistry; D008555:Melitten / Q000494:pharmacology*; D019301:Oleic Acid / Q000378:metabolism*; D014357:Trypsin / Q000737:chemistry*
D007202:Indicators and Reagents; D008055:Lipids; D008555:Melitten; D019301:Oleic Acid; D014357:Trypsin