title: A protein network for phospholipid synthesis uncovered by a variant of the tandem affinity purification method in Escherichia coli.
abstract: In prokaryotes, acyl carrier protein (ACP) is a cofactor central to a myriad of syntheses, including fatty acid and phospholipid synthesis. To fulfill its function, ACP must therefore interact with a multitude of different enzymes, which includes the thioesterase YbgC. We found a specific interaction between ACP and YbgC whose thioesterase activity has been demonstrated in vitro on acyl-CoA derivatives, but whose physiological function in bacteria remains unknown. Therefore, YbgC could be a thioesterase active on some specific acyl-ACPs. We then assigned a function to the ACP/YbgC pair by employing a proteomic approach derived from tandem affinity purification, the split tag method. This technique allowed us to purify proteins interacting with ACP and YbgC proteins at the same time. Interactions with PlsB, a sn-glycerol-3-phosphate acyltransferase and PssA, a phosphatidylserine synthase, were identified and validated, showing that YbgC is involved in phospholipid metabolism. Furthermore, using an in vivo bacterial two-hybrid interaction analysis, we showed for the first time that enzymes of the phospholipid synthesis pathway form a complex in the inner membrane. Taken together, these results describe an integrated protein network that could be involved in the coordination of phospholipid metabolism.
authors.affiliation: LISM, IBSM, CNRS, 31 chemin Joseph Aiguier, 13402 Marseille, France.
authors.forename: Djamel
authors.initials: D
authors.lastname: Gully
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Bouveret
mesh_terms: D015153:Blotting, Western; D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000302:isolation & purification / Q000378:metabolism*; D010743:Phospholipids / Q000138:chemical synthesis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D029968:Escherichia coli Proteins; D010743:Phospholipids