Functional characterization of human 1-acylglycerol-3-phosphate-O-acyltransferase isoform 9: cloning, tissue distribution, gene structure, and enzymatic activity

Abstract

Most cells synthesize their glycerophospholipids and triglycerides (TG) to maintain the cellular integrity and to provide energy for cellular functions. The phospholipids are synthesized de novo in cells through an evolutionary conserved process involving serial acylations of glycerol-3-phosphate. Several isoforms of the enzyme 1-acylglycerol-3-phosphate-O-acyltransferase (AGPAT) acylate lysophosphatidic acid at the sn-2 position to produce phosphatidic acid. We cloned a cDNA predicted to be an AGPAT isoform and designated it AGPAT9. The human AGPAT9 gene spans across 14 exons and encodes for a polypeptide of 534 amino acids. AGPAT9 is highly expressed in the lung and spleen, followed by leukocyte, omental adipose tissue, and placenta. In the Chinese Hamster Ovary (CHO), cell lysates overexpressing AGPAT9, we observed AGPAT activity but not the lysophosphatidylcholine acyltransferase activity. When AGPAT9 is coexpressed with AGPAT1 in CHO cells, both the isoforms localize to the endoplasmic reticulum (ER) and occupy the same ER domain as AGPAT1. Despite substitution of asparagine with proline in the NHX 4 D motif and arginine with cysteine in the EGTR motif, AGPAT9 retains AGPAT activity suggesting that residues asparagine and arginine in the NHX 4 D and EGTR motifs respectively are not essential for the enzymatic activity. Based on the X-ray crystallographic structure of a related acyltransferase, squash gpat, a model is proposed in which a hydrophobic pocket in AGPAT9 accommodates fatty acyl chains of both substrates in an orientation, whereas the HX 4 D motif participates in catalysis. Based on the activity and expression pattern of AGPAT9 in the lung and spleen, this novel isoform could be implicated in the biosynthesis of phospholipids and TG in these tissues.

Molecular cloning, sequence characterization, and gene expression profiling of a novel water buffalo (Bubalus bubalis) gene, AGPAT6

“…AGPAT6 is broadly expressed and can be detected at the mRNA level in multiple tissues examined Agarwal et al, 2007;Chen et al, 2008). In mouse, AGPAT6 is expressed at a high level in brown adipose tissue, white adipose tissue, liver, and mammary epithelium of breast tissue Bionaz and Loor, 2008a).…”

“…It catalyzes the second step by acylating lysophosphatidic acid to phosphatidic acid (Aguado and Campbell, 1998;Coleman and Lee, 2004;Ye et al, 2005;Agarwal et al, 2006Agarwal et al, , 2007Nagle et al, 2008;Sukumaran et al, 2009). So far, eight members of the AGPAT gene family have been described in humans, which are AGPAT1-8, and each of them possesses a lysophosphatidic acid acyltransferase motif (Ye et al, 2005;Nagle et al, 2008).…”

AGPAT6 Is a Novel Microsomal Glycerol-3-phosphate Acyltransferase

“…Subcellular Localization and the Tissue Distribution of Human GPAT4-Mouse AGPAT6 was shown to be located within the endoplasmic reticulum (20), but differences between mouse and human glycerolipid acyltransferases have been reported (17,18,27). Thus, we performed experiments to define the location of human AGPAT6 (GPAT4).…”

Pathology of Congenital Generalized Lipodystrophy in Agpat2–/– Mice

“…35 The AGPAT2 enzyme represents 1 of at least 10 isoforms of AGPAT, all of which have different tissue expression profiles. 2,3,5,20,36,47,48 The AGPAT2 enzyme (also known as lysophosphatidic acid acyltransferase) is most highly expressed in adipose tissue, 2 where it catalyzes the acylation of lysophosphatidic acid to phosphatidic acid (which is an essential precursor in the biosynthesis of triacylglycerol and phospholipids). 30,32 Mouse models with lipid disorders have provided important insights into the pathogenetic role of adipocytes in the development of insulin resistance and diabetes and in the processes of adipocyte differentiation.…”