The Chemistry of Phospholipid Biosynthesis
Phospholipids are the major constituents of the cell membrane, the water-impermeable barrier that surrounds each and every cell, crucial for its existence, wellbeing and survival. All cells, prokaryotic and eukaryotic alike, need to be able to assemble their membranes, and synthesize their components. The key step in phospholipid biosynthesis involves the reaction of a diacylglycerol lipid tail with the polar group that gives the identity – for example choline, inositol, and ethanolamine to form phosphatidylcholine, phosphatidylinositol and phosphatidylethanolamine respectively. This key step is catalyzed by a large family of enzymes called CDP-alcohol phosphotransferases (CDP-APs). The reaction involves the transfer of a CDP-activated donor alcohol group to an acceptor alcohol, as for example CDP-attached diacylglycerol to choline, to form phosphatidylcholine. CDP-APs are all integral membrane proteins, characterized by a universally conserved signature motif of eight amino acids spaced over a small stretch of sequence (D1xxD2G1xxAR…G2xxxD3xxxD4). The absolute conservation of this motif in enzymes with such diverse substrate specificities suggests that it plays a fundamental role in catalysis, but the lack of any structural information has hampered progress in understanding the mechanism of this essential class of enzymes.
The structure of AF2299 shows what to the best of the author’s knowledge is a novel fold consisting of 6 transmembrane (TM) segments surrounding a polar cavity. This cavity spans from deep within the membrane to the cytosol, and constitutes the active site. The signature sequence, split between TM2 and TM3 lines the active site. Structures with and without CDP and the CDP-bound donor substrate, as well as with the bound product CMP, allow the authors to assign an unambiguous role to each of the 8 conserved residues in the signature sequence. The AF2299 structures reveal that D1, D2 and D3 coordinate the pyro-phosphate of CDP via a divalent cation. In contrast the authors suggest that D4 acts as a catalytic base, abstracting a proton from the terminal hydroxyl of the acceptor alcohol, facilitating nucleophilic attack by the activated acceptor on the β-phosphorus of the CDP. The structures also show the donor and acceptor substituent binding pockets. Given the absolute geometrical conservation of the key elements within the active site, the position of the donor and acceptor substrates will most likely also remain constant, so as to allow catalysis. Therefore, the structures reported here will also provide a framework in which to investigate the molecular elements underlying substrate specificity for CDP-APs. This research was supported by the National Institutes of Health (NIH) grant GM095315
Nature Communications, June 2014