COMPLETE DISCRIMINATION OF DOCOSAHEXAENOATE FROM ARACHIDONATE BY 85 KDA CYTOSOLIC PHOSPHOLIPASE A(2) DURING THE HYDROLYSIS OF DIACYL AND ALKENYLACYLGLYCEROPHOSPHOETHANOLAMINE
M. Shikano et al., COMPLETE DISCRIMINATION OF DOCOSAHEXAENOATE FROM ARACHIDONATE BY 85 KDA CYTOSOLIC PHOSPHOLIPASE A(2) DURING THE HYDROLYSIS OF DIACYL AND ALKENYLACYLGLYCEROPHOSPHOETHANOLAMINE, Biochimica et biophysica acta, L. Lipids and lipid metabolism, 1212(2), 1994, pp. 211-216
In our previous report (Shikano, M., Masuzawa, Y. and Yazawa, K. (1993
) J. Immunol. 150, 3525-3533), we described that the enrichment of doc
osahexaenoic acid (DHA, 22:6(n - 3)) reduces both arachidonic acid (AA
, 20:4(n - 6)) release and platelet-activating factor (PAF) synthesis
in human eosinophilic leukemia cells, Eol-1. Since no DHA release was
observed in response to Ca-ionophore stimulation, we presumed that the
phospholipase A(2) (PLA(2)) responsible for AA release and PAF synthe
sis can not hydrolyze the DHA moiety of phospholipids. In the present
paper, we examined whether DHA-containing diacyl- and alkenylacylglyce
rophosphoethanolamine (DHA-diacylGPE and DHA-alkenylacyGPE) are suscep
tible to the action of AA-preferential 85 kDa cytosolic phospholipase
A(2) (cPLA(2)) from rabbit platelets in comparison with AA and eicosap
entaenoic acid (EPA, 20:5(n - 3)) derivatives. When diacylGPE was used
as a substrate, DHA release was almost negligible under the assay con
dition that allowed AA and EPA to be liberated at the rates of 4.3 mu
mol/min per mg protein and 2.5 mu mol/min per mg protein, respectively
. On the other hand, 14 kDa type II PLA, hydrolyzed DHA-diacylGPE as w
ell as AA-diacylGPE and EPA-diacylGPE. When DHA-diacylGPE and AA-diacy
lGPE were mixed at equimolar concentrations, DHA release by cPLA(2) wa
s not observed and AA release was reduced to 32% in the case without D
HA-diacylGPE. This indicated that DHA-diacylGPE is a poor substrate bu
t possesses the inhibitory activity for cPLA(2). cPLA(2) does not clea
rly discriminate between AA-alkenylacylGPE and AA-diacylGPE. As in the
case using diacylGPE as a substrate, DHA-alkenylacylGPE was completel
y discriminated from AA-alkenylacylGPE by cPLA(2). The roles of DHA an
d cPLA(2) in the synthesis of lipid mediators will be discussed in rel
ation to the new aspects of the substrate specificity of cPLA(2) provi
ded here.