title: Cloning and functional characterization of a novel mitochondrial N-ethylmaleimide-sensitive glycerol-3-phosphate acyltransferase (GPAT2).
abstract: Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step in glycerolipid synthesis. Several mammalian GPAT activities have been recognized, including N-ethylmaleimide (NEM)-sensitive isoforms in microsomes and mitochondria and an NEM-resistant form in mitochondrial outer membrane (GPAT1). We have now cloned a second mitochondrial isoform, GPAT2 from mouse testis. The open-reading frame encodes a protein of 798 amino acids with a calculated mass of 88.8kDa and 27% amino acid identity to GPAT1. Testis mRNA expression was 50-fold higher than in liver or brown adipose tissue, but the specific activity of NEM-sensitive GPAT in testis mitochondria was similar to that in liver. When Cos-7 cells were transiently transfected with GPAT2, NEM-sensitive GPAT activity increased 30%. Confocal microscopy confirmed a mitochondrial location. Incubation of GPAT2-transfected Cos-7 cells with trace (3 microM; 0.25 microCi) [1-(14)C]oleate for 6h increased incorporation of [(14)C]oleate into TAG 84%. In contrast, incorporation into phospholipid species was lower than in control cells. Although a polyclonal antibody raised against full-length GPAT1 detected an approximately 89-kDa band in liver and testis from GPAT1 null mice and both 89- and 80-kDa bands in BAT from the knockout animals, the GPAT2 protein expressed in Cos-7 cells was only 80 kDa. In vitro translation showed a single product of 89 kDa. Unlike GPAT1, GPAT2 mRNA abundance in liver was not altered by fasting or refeeding. GPAT2 is likely to have a specialized function in testis.
authors: Department of Nutrition, University of North Carolina-Chapel Hill, Chapel Hill, NC 27599, USA.
authors: Shuli
authors: S
authors: Wang
authors: Douglas P
authors: DP
authors: Lee
authors: Nan
authors: N
authors: Gong
authors: Nicole M J
authors: NM
authors: Schwerbrock
authors: Douglas G
authors: DG
authors: Mashek
authors: Maria R
authors: MR
authors: Gonzalez-Baró
authors: Cliona
authors: C
authors: Stapleton
authors: Lei O
authors: LO
authors: Li
authors: Tal M
authors: TM
authors: Lewin
authors: Rosalind A
authors: RA
authors: Coleman
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular / Q000379:methods; D004789:Enzyme Activation; D005992:Glycerol-3-Phosphate O-Acyltransferase / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008928:Mitochondria / Q000201:enzymology*; D009928:Organ Specificity; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013737:Testis / Q000201:enzymology*; D014018:Tissue Distribution
chemical_list: D011994:Recombinant Proteins; D005992:Glycerol-3-Phosphate O-Acyltransferase
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