Glycolipids isolated from Aplysia kurodai can activate cyclic adenosine 3',5'-monophosphate-dependent protein kinase from rat brain.

Abstract

Cyclic AMP (cAMP)-dependent protein kinase (cAMP-kinase) partially purified from the membrane fractions of rat brains was stimulated by novel phosphonoglycosphingolipids (glycolipids) derived from the skin and nerve fibers of Aplysia kurodai. Among various glycolipids tested, a major glycolipid from the skin, 3-O-MeGal beta 1-->3GalNAc alpha 1-->3[6'-O-(2-aminoethylphosphonyl)Gal alpha 1-->2](2-aminoethylphosphonyl-->6)Glc beta 1-->4Glc beta 1-->1ceramide (SGL-II), was most potent, giving half-maximal activation at 32.2 microM. Activation of cAMP-kinase was maximal with 250 microM SGL-II using kemptide as substrate. The effect of SGL-II was additive on kinase activity at submaximal concentrations of cAMP. The kinase activity activated with SGL-II was inhibited by the addition of protein kinase inhibitor peptide, a specific peptide inhibitor for cAMP-kinase. Its inhibitory pattern was similar to that for the catalytic subunit. Of the various substrates tested, the glycolipid-stimulated cAMP-kinase could phosphorylate microtubule-associated protein 2, synapsin I, and myelin basic protein but not histone H1 and casein. The regulatory subunit strongly inhibited the activity of purified catalytic subunit of cAMP-kinase. This inhibition was reversed by addition of SGL-II, as observed for cAMP. SGL-II was capable of partially dissociating cAMP-kinase, which was observed by gel filtration column chromatography. However, the binding activity of cAMP to the holoenzyme was not inhibited with SGL-II. These results demonstrate that the glycolipids can directly activate cAMP-kinase in a manner similar, but not identical, to that of cAMP.