Diacylglycerol kinase epsilon is selective for both acyl chains of phosphatidic acid or diacylglycerol.
The phosphatidylinositol (PI) cycle mediates many cellular events by controlling the metabolism of many lipid second messengers. Diacylglycerol kinase epsilon (DGK epsilon) has an important role in this cycle. DGK epsilon is the only DGK isoform to show inhibition by its product phosphatidic acid (PA) as well as substrate specificity for sn-2 arachidonoyl-diacylglycerol (DAG). Here, we show that this inhibition and substrate specificity are both determined by selectivity for a combination of the sn-1 and sn-2 acyl chains of PA or DAG, respectively, preferring the most prevalent acyl chain composition of lipids involved specifically in the PI cycle, 1-stearoyl-2-arachidonoyl. Although the difference in rate for closely related lipid species is small, there is a significant enrichment of 1-stearoyl-2-arachidonoyl PI because of the cyclical nature of PI turnover. We also show that the inhibition of DGK epsilon by PA is competitive and that the deletion of the hydrophobic segment and cationic cluster of DGK epsilon does not affect its selectivity for the acyl chains of PA or DAG. Thus, this active site not only recognizes the lipid headgroup but also a combination of the two acyl chains in PA or DAG. We propose a mechanism of DGK epsilon regulation where its dual acyl chain selectivity is used to negatively regulate its enzymatic activity in a manner that ensures DGK epsilon remains committed to the PI turnover cycle. This novel mechanism of enzyme regulation within a signaling pathway could serve as a template for the regulation of enzymes in other pathways in the cell.
Lung
Michael
M
Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
Shulga
Yulia V
YV
Ivanova
Pavlina T
PT
Myers
David S
DS
Milne
Stephen B
SB
Brown
H Alex
HA
Topham
Matthew K
MK
Epand
Richard M
RM
D000818:Animals; D019556:COS Cells; D020134:Catalytic Domain; D002522:Chlorocebus aethiops; D019852:Diacylglycerol Kinase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004075:Diglycerides / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D015394:Molecular Structure; D010712:Phosphatidic Acids / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D004075:Diglycerides; D010712:Phosphatidic Acids; D011994:Recombinant Proteins; C526464:DGKE protein, human; D019852:Diacylglycerol Kinase