Purification and characterization of two phospholipid exchange proteins from bovine heart.
Two phospholipid exchange proteins from bovine heart have been purified approximately 2000-fold and judged greater than 90% pure. The proteins are similar in molecular weight (both 33,400 by polyacrylamide gel electrophoresis and 23,500 by gel filtration), in amino acid composition, and in specificity, although they differ in isoelectric points, 5.3 and 5.6. The transfer of phospholipids between artificial membranes is catalyzed by these proteins at the following relative rates: 100 for phosphatidylinositol, 35 for phosphatidylcholine, 5 for sphingomyelin, and 0.1 for phosphatidylethanolamine. The use of these exchange proteins in the study of mixed phospholipid vesicle structure is demonstrated. The purified proteins catalyze the substitution of one membrane phospholipid species for another at a rate comparable to true exchange. The phospholipid exchange activity is inhibited by the presence of sphingomyelin, and also by reagents which react with sulfhydryl groups. Evidence is presented for two sites of N-ethylmaleimide binding on these exchange proteins. Reaction with one site has little effect on activity and occurs in the absence of membranes. Reaction with the second site occurs in the presence of phospholipid vesicles and leads to complete, irreversible inhibition of exchange activity.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D005033:Ethylmaleimide / Q000494:pharmacology; D007700:Kinetics; D008567:Membranes, Artificial; D008970:Molecular Weight; D009206:Myocardium / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D000596:Amino Acids; D002352:Carrier Proteins; D008567:Membranes, Artificial; D010743:Phospholipids; D013439:Sulfhydryl Reagents; D005033:Ethylmaleimide