Glycolipids isolated from Aplysia kurodai can activate cyclic adenosine 3',5'-monophosphate-dependent protein kinase from rat brain.
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.
Arakane
F
F
Department of Pharmacology, Kumamoto University School of Medicine, Japan.
Fukunaga
K
K
Araki
S
S
Abe
S
S
Satake
M
M
Miyazaki
K
K
Okamura
H
H
Miyamoto
E
E
D000818:Animals; D001048:Aplysia; D001921:Brain / Q000201:enzymology*; D002240:Carbohydrate Sequence; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation; D006017:Glycolipids / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000302:isolation & purification; D051381:Rats; D012867:Skin; D013379:Substrate Specificity; D014461:Ultracentrifugation
D006017:Glycolipids; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D009844:Oligosaccharides; D017868:Cyclic AMP-Dependent Protein Kinases