title: Solubilisation, partial purification and properties of acyl-CoA: glycerol-3-phosphate acyltransferase from avocado (Persea americana) fruit mesocarp.
abstract: Glycerol-3-phosphate acyltransferase (G3PAT) activity was studied using a microsomal membrane fraction from avocado (Persea americana) mesocarp. G3PAT was shown to be an integral membrane protein, having an active site that appeared to be accessible to the cytoplasmic face of the endoplasmic reticulum, in experiments using limited proteolytic digestion. CHAPS solubilisation (0.25%, w/v) of microsomal G3PAT activity was used as an initial step in purification of this enzyme. Both CHAPS-solubilised and microsomal G3PAT activities were characterised and compared. Affinity chromatography was used to purify microsomal G3PAT for the first time from a plant source. Glycerophosphorylethanolamine coupled to cyanogen bromide-activated Sepharose was used for this purpose. Specific elution of G3PAT by a solution of glycerol-3-phosphate resulted in about 150-fold purification. The significance of the results and the potential usefulness of the purification method for further studies of G3PAT in plants are discussed.
authors.affiliation: Department of Biochemistry, University of Wales College of Cardiff, UK.
authors.forename: V S
authors.initials: VS
authors.lastname: Eccleston
authors.forename: J L
authors.initials: JL
authors.lastname: Harwood
mesh_terms: D001665:Binding Sites; D002793:Cholic Acids; D002846:Chromatography, Affinity; D003902:Detergents; D005638:Fruit / Q000201:enzymology*; D005992:Glycerol-3-Phosphate O-Acyltransferase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008861:Microsomes / Q000201:enzymology; D012995:Solubility
chemical_list: D002793:Cholic Acids; D003902:Detergents; D005992:Glycerol-3-Phosphate O-Acyltransferase; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate