Flippability of Escherichia coli membrane proteins under lipid stress reveals a new job for old genes

Normally, the “flip-flop” motion is reserved for small lipid molecules which move from one leaflet of the membrane to the other, and even that process is very slow without the aid of specific enzymes. Unexpectedly, we recently found that despite a prohibitively high energy barrier, Escherichia coli cardiolipin (CL) synthase A (ClsA) flips its large catalytic domain across the inner (cytoplasmic) membrane (IM) when cells were fully or partially depleted of phosphatidylethanolamine (PE) and that this was necessary to support cell viability. Here, we demonstrated that production of the non-native phospholipid phosphatidylcholine (PC) in PE-depleted cells prevents ClsA flipping and halts cell growth, a phenotype rescued by constitutively inverted ClsA. Surprisingly, properly oriented lactose permease (LacY) acts as a multicopy suppressor, rescuing the growth defect in the -PE+PC cells. These data indicate that neutral PC regulates the membrane topology of ClsA and LacY, and subsequently the appearance of CL to the periplasmic leaflet required for viability. Therefore, ClsA and potentially LacY may achieve varied roles from the same amino acid sequence through an unusual flip-flopping mechanism. This fruitful “repurposing” of genetic material highlights the remarkable adaptability of living organisms and their membranes. Altering E. coli lipids disrupts growth, but controlled membrane protein flipping simulates natural selection and resemonstrates that membrane proteins can re-evolve under the selective pressure of an altered lipid bilayer.