Adenylyl cyclase in yeast. Hydrodynamic properties and activation by trypsin.
The adenylyl cyclase system of the yeast Saccharomyces cerevisiae contains the CYR1 polypeptide, responsible for catalyzing formation of cAMP from ATP, and two RAS polypeptides, responsible for stimulation of cAMP synthesis by guanine nucleotides. We have determined hydrodynamic properties of yeast adenylyl cyclase in taurocholate extracts of wild type and RAS-deficient membranes. In taurocholate extracts of both kinds of membranes, the enzyme is insensitive to guanine nucleotide stimulation; in the presence of 0.5 M NaCl, the taurocholate-solubilized enzyme has a sedimentation coefficient of 12.5 S and a Stokes radius of 11 nm, consistent with a molecular weight of 594,000 for the protein-detergent complex. Treatment of particulate fractions with trypsin (less than 10 micrograms/ml) markedly activates membrane-bound adenylyl cyclase activity, abolishes stimulation by guanine nucleotides, and reduces the sedimentation coefficient of the detergent-solubilized enzyme; higher concentrations of trypsin release a still smaller water-soluble enzyme complex (7.5 S, 6.1 nm Stokes radius, calculated Mr = 190,000) from the membrane. In combination with genetic evidence (Kataoka, T., Broek, D., and Wigler M., (1985) Cell 43, 493-505), our data are consistent with a structural and functional model of yeast adenylyl cyclase in which GTP-activated RAS proteins stimulate cAMP synthesis by relieving an inhibitory constraint on the activity of the CYR1 gene product. This constraint may be mediated by the amino-terminal portion of the CYR1 polypeptide.
D000255:Adenosine Triphosphate / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D002498:Centrifugation; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D000242:Cyclic AMP / Q000096:biosynthesis; D004789:Enzyme Activation / Q000187:drug effects; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology; D006150:Guanine Nucleotides / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D012965:Sodium Chloride / Q000494:pharmacology; D013656:Taurocholic Acid; D014357:Trypsin / Q000494:pharmacology; D018631:ras Proteins
D005656:Fungal Proteins; D006150:Guanine Nucleotides; D008345:Manganese; D012965:Sodium Chloride; D013656:Taurocholic Acid; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D014357:Trypsin; D018631:ras Proteins; D000262:Adenylyl Cyclases; D008274:Magnesium