FATTY-ACID FLIP-FLOP IN PHOSPHOLIPID-BILAYERS IS EXTREMELY FAST

The rate of movement of fatty acids (FA) across phospholipid bilayers is an important consideration for their mechanism of transport across cell membranes but has not yet been measured. When FA move undirection ally across phospholipid bilayers, the rapid movement of un-ionized FA compared to ionized FA results in transport of protons. We have previ ously used this property to show that FA move spontaneously (''flip-fl op'') across the bilayer of small unilamellar vesicles within similar to 1 s (Kamp & Hamilton, 1992, 1993). This work extends the time resol ution of this assay into the millisecond time range by use of stopped flow fluorometry. In small unilamellar vesicles (diameter, similar to 25 nm) at neutral pH, flip-flop of all fatty acids studied (lauric, my ristic, palmitic, oleic, and stearic) was greater than or equal to 80% complete within 5-10 ms. In large unilamellar vesicles (diameter, sim ilar to 100 nm), the same fatty acids exhibited fast flip-flop but wit h a measureable rate (t(1/2) = 23 +/- 12 ms). The calculated pseudouni molecular rate constant of the un-ionized FA (k(FAH)) similar to 15 S- 1. There was no dependence of the flip-flop rate on the fatty acid cha in length or structure. We also monitored the rate of desorption and t ransbilayer movement of (anthroyloxy)stearic acid in small unilamellar vesicles. Whereas previous studies suggested slow flip-flop of this F A analogue, the present studies suggest that (anthroyloxy)stearic acid flip-flops rapidly and that earlier studies did not truly measure the transbilayer movement step. These findings further support the view t hat proteins are not required for translocation of FA across cell memb ranes.