Abstract

BACKGROUND:

'Giant vesicles' are liposomes that have diameters of several micrometers. It is possible to microinject biochemicals into a single vesicle and follow the progress of a chemical reaction in real time by light microscopy. We have previously used this technique to inject phospholipase A2 into giant vesicles; the vesicles disappeared as their 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-glycerol-3-phosphate. Microinjection of the enzyme sn-glycerol-3-phosphate-acyltransferase into the vesicle catalyzes the in situ production of the lipid membrane precursor 1-palmitoyl-sn-glycerol-3-phosphate, which remains incorporated in the membrane. The altered membrane chemistry 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.

CONCLUSIONS:

We have used the first step of the 'salvage pathway' for synthesis of POPC to demonstrate that it is possible to localize the synthesis of a lipid membrane precursor inside a giant vesicle. In the future it may be possible to combine the necessary enzymes and substrates to carry out the reactions for a complete metabolic pathway within a liposome.