ENZYME-CONTAINING LIPOSOMES CAN ENDOGENOUSLY PRODUCE MEMBRANE-CONSTITUTING LIPIDS

Background: 'Giant vesicles' are liposomes that have diameters of seve ral micrometers. It is possible to microinject biochemicals into a sin gle vesicle and follow the progress of a chemical reaction in real tim e by light microscopy. We have previously used this technique to injec t phospholipase A:! into giant vesicles; the vesicles disappeared as t heir components were hydrolyzed. Here we investigate whether the lipid components of a vesicle can be synthesized inside it. Results: Giant vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine (POPC) and palmitoyl-CoA were prepared in a solution containing sn-gl ycerol-3-phosphate. Microinjection of the enzyme sn-glycerol-3-phospha te-acyltransferase into the vesicle catalyzes the in situ production o f the lipid membrane precursor 1-palmitoyl-sn-glycerol-3-phosphate, wh ich remains incorporated in the membrane. The altered membrane chemist ry causes shrinkage of the vesicle and formation of smaller liposomes on the inner surface at the site of injection. Similar transformations were seen when the enzyme was added to the outside of the vesicle. Co nclusions: We have used the first step of the 'salvage pathway' for sy nthesis of POPC to demonstrate that it is possible to localize the syn thesis of a lipid membrane precursor inside a giant vesicle. In the fu ture it may be possible to combine the necessary enzymes and substrate s to carry out the reactions for a complete metabolic pathway within a liposome.