Substrate chirality and specificity of diacylglycerol kinases and the multisubstrate lipid kinase.
The alpha, zeta, and epsilon isoforms of diacylglycerol kinase exhibit a high degree of stereospecificity in the phosphorylation of diacylglycerol. In comparison, a multiple lipid kinase, MuLK, shows much less stereospecificity, phosphorylating 1,2-dioleoylglycerol only approximately 2-3 times more rapidly than 2,3-dioleoylglycerol. The alpha and zeta isoforms of diacylglycerol kinase are inhibited by 2,3-dioleoylglycerol, but not the more substrate-selective epsilon isoform. The inhibition by 2,3-dioleoylglycerol is uncompetitive. This corresponds to a kinetic scheme in which the inhibitor can bind to the enzyme-substrate complex, but not to the free enzyme. Our data indicate that despite their similar structures, 1,2-dioleoylglycerol and 2,3-dioleoylglycerol do not compete for the active site of these three isoforms of diacylglycerol kinase. We suggest that the 2,3-dioleoylglycerol binds to a site on the alpha and zeta isoforms of diacylglycerol kinase that is exposed as a consequence of the substrate binding to the active site. The chiral specificity of these enzymes thus mimics the substrate specificity, with MuLK being the least selective and the epsilon isoform of diacylglycerol kinase exhibiting the greatest selectivity.
Epand
Richard M
RM
Department of Biochemistry and Biomedical Sciences, McMaster University, 1200 Main Street West, Hamilton, Ontario L8N 3Z5, Canada. epand@mcmaster.ca
Shulga
Yulia V
YV
Timmons
Heath C
HC
Perri
Alexandra L
AL
Belani
Jitendra D
JD
Perinpanathan
Kirishanth
K
Johnson-McIntire
Laura Beth
LB
Bajjalieh
Sandra
S
Dicu
Armela O
AO
Elias
Cynthia
C
Rychnovsky
Scott D
SD
Topham
Matthew K
MK
D001665:Binding Sites; D019852:Diacylglycerol Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004075:Diglycerides / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008071:Lipoprotein Lipase / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D013237:Stereoisomerism; D013379:Substrate Specificity
D004075:Diglycerides; C492429:AGK protein, human; D017853:Phosphotransferases (Alcohol Group Acceptor); C526464:DGKE protein, human; C526463:DGKZ protein, human; D019852:Diacylglycerol Kinase; C480280:DAGLA protein, human; D008071:Lipoprotein Lipase; C013965:diolein