Trypsin-induced lysis of lipid vesicles: effect of surface charge and lipid composition.
We have made a curious observation that the proteolytic enzyme, trypsin, induced a rapid and complete release of the contents of vesicles composed of dioleoylphosphatidylethanolamine (DOPE) and oleic acid (OA). Content release at 37 degrees C, monitored by the release of an entrapped fluorescence marker (calcein), was accompanied by an extensive vesicle aggregation. The lytic activity of trypsin on the vesicles depended on pH and liposome composition. The optimal pH for vesicle lysis was below pH 7.4, which was different from the optimal pH for catalytic activity of trypsin. The lytic activity of trypsin was specific for vesicles composed of DOPE and fatty acids such as OA and palmitoleic acid; vesicles composed of dioleoylphosphatidylcholine, N-methyl-DOPE, and OA, or DOPE combined with other negatively charged lipids such as phosphatidylserine and phosphatidic acid were not sensitive to trypsin. Inhibition of enzyme activity by trypsin inhibitors did not abolish the lytic activity, suggesting that the lytic activity of trypsin is not related to the catalytic activity. However, the lytic activity of trypsin on vesicles composed of DOPE and OA was inhibited in the presence of excess vesicles containing negative charges, or by a pretreatment of trypsin with acylating reagent to reduce the positive-charge content of trypsin. These data demonstrate that vesicle aggregation and lysis are the results of electrostatic interactions of positive charges on trypsin and negative charges on the vesicles. Phase separation and transition to nonbilayer phases of the vesicle lipids are likely involved.
Liu
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D
Department of Biochemistry, University of Tennessee, Knoxville 37996-0840.
Huang
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L
D005452:Fluoresceins / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D008081:Liposomes / Q000737:chemistry*; D019301:Oleic Acid; D009829:Oleic Acids / Q000737:chemistry*; D010714:Phosphatidylethanolamines / Q000737:chemistry*; D010743:Phospholipids / Q000737:chemistry*; D013499:Surface Properties; D014357:Trypsin / Q000737:chemistry / Q000494:pharmacology*
D005452:Fluoresceins; D008081:Liposomes; D009829:Oleic Acids; D010714:Phosphatidylethanolamines; D010743:Phospholipids; D019301:Oleic Acid; C020888:1,2-dielaidoylphosphatidylethanolamine; D014357:Trypsin; C007740:fluorexon