title: Characterization of active and latent forms of the membrane-associated sn-glycerol-3-phosphate acyltransferase of Escherichia coli.
abstract: The intrinsically active, sn-glycerol-3-phosphate acyltransferase present in membranes prepared from both wild type Escherichia coli and from strains which overproduce the enzyme can be kinetically distinguished from a latent enzyme species which is unmasked by solubilization and reconstitution. Both membrane-associated and solubilized/reconstituted enzyme preparations exhibited cooperativity with respect to sn-glycerol-3-phosphate and palmitoyl-coenzyme A substrates; positive cooperativity in membranes toward palmitoyl-coenzyme A (napp = 4) and negative cooperativity toward sn-glycerol-3-phosphate (napp = 0.75) were significantly altered upon solubilization and reconstitution. Since the degree of alteration increased with the amount of sn-glycerol-3-P acyltransferase present in the membranes, a detergent-dissociable homooligomerization of the sn-glycerol-3-phosphate acyltransferase was considered as an underlying mechanism. This possibility was investigated by changing the protein-to-Triton X-100 ratio of homogeneous enzyme prior to reconstitution and then analyzing the subsequent migration of samples on a Sephacryl S-300 sizing column. The elution positions were consistent with monomeric and dimeric polypeptide bound to micelles of Triton X-100. Hill coefficients for monomeric, reconstituted enzyme preparations were comparable to those obtained for the active, membrane-associated sn-glycerol-3-phosphate acyltransferase. The reduced cooperativity of dimeric, reconstituted enzyme preparations correlated closely to the Hill coefficient values obtained for latent, solubilized/reconstituted sn-glycerol-3-phosphate acyltransferase from membranes of Escherichia coli which overproduce the enzyme. The physiological significance of these findings is discussed.
authors.affiliation: Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710.
authors.forename: M A
authors.initials: MA
authors.lastname: Scheideler
authors.forename: R M
authors.initials: RM
authors.lastname: Bell
mesh_terms: D002462:Cell Membrane / Q000201:enzymology; D002850:Chromatography, Gel; D004926:Escherichia coli / Q000201:enzymology*; D005992:Glycerol-3-Phosphate O-Acyltransferase / Q000378:metabolism*; D007700:Kinetics; D008823:Micelles; D017830:Octoxynol; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D011092:Polyethylene Glycols; D013379:Substrate Specificity; D013501:Surface-Active Agents
chemical_list: D008823:Micelles; D013501:Surface-Active Agents; D010171:Palmitoyl Coenzyme A; D011092:Polyethylene Glycols; D017830:Octoxynol; D005992:Glycerol-3-Phosphate O-Acyltransferase