Structure of brain adenylate cyclase: proteolysis-dependent modifications.
The associations of the components of eucaryotic adenylate cyclase are still poorly characterized. Enzyme activity is, however, thought to depend upon subunit conformations and states of association. Estimates of adenylate cyclase sizes corresponding to given levels of activity may thus give clues as to how the enzyme functions. Studying the rat brain enzyme, we found that samples protected from proteolysis throughout the fractionation procedure yielded, upon Lubrol solubilization, a soluble protein complex of 9.1S sedimentation coefficient and 11.5-nm Stokes radius. These values are much larger than those previously reported. The soluble enzyme specific activity, but not its size, was dependent upon the various effectors preincubated with the membranes. Proteolysis is known to first activate and then decrease adenylate cyclase activity. Proteolysis of the brain samples, whether due to trypsin or to endogeneous proteases, decreased the adenylate cyclase s value, Stokes radius, and specific activity altogether. The magnitude of the shifts depended upon the nature of the enzyme effector preincubated with the membranes. We recently showed that some brain membrane proteins can be ADP-ribosylated by cholera toxin, concomitantly with adenylate cyclase activation [Berthillier, G., d'-Alayer, J., & Monneron, A. (1982) Biochem. Biophys. Res. Commun. 109, 297-304]. Trypsin treatment of such samples led to a quick degradation of the labeled polypeptides and especially of the Mr 47000 protein. This Lubrol soluble protein is likely to be the brain G/F stimulatory subunit.
D000262:Adenylyl Cyclases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002712:Chlorides; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D008345:Manganese / Q000494:pharmacology; D017895:Manganese Compounds; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains; D013570:Synaptic Membranes / Q000201:enzymology; D013574:Synaptosomes / Q000201:enzymology*
D002712:Chlorides; D017895:Manganese Compounds; D015636:Magnesium Chloride; D008345:Manganese; D000262:Adenylyl Cyclases; D008274:Magnesium; C025340:manganese chloride