Phospholipid and guanine nucleotides sensitive properties of the smooth muscle adenylate cyclase catalytic unit.
The adenylate cyclase catalytic unit was partially purified from uterine smooth muscle by chromatography on columns of SM-2 Bio-Beads and Sepharose 6B. Stimulation of catalysis by forskolin was much greater in the presence of Mn2+ than in the presence of Mg2+. Neither NaF nor guanine nucleotide stimulated catalysis in the presence of Mg2+ or Mn2+. These properties indicated the catalytic unit was not sensitive to regulation by the GS regulatory protein. Guanine nucleotide inhibited catalysis, however, and was a competitive inhibitor of the ATP substrate (Ki approximately 50 microM). Since inhibition affected Km but not Vmax, the catalytic unit also seemed insensitive to regulation by the Gi regulatory protein, which does not act like a competitive inhibitor in other enzyme systems. The catalytic unit was also phospholipid sensitive. Only phosphatidic acid (Pho-A) had a direct effect on catalysis and was a potent inhibitor. Its effects were antagonized by the concomitant addition of phosphatidylcholine (Pho-C) but not by phosphatidylethanolamine, phosphatidylserine, or phosphatidylinositol. Acyl chain composition had a marked effect on Pho-C binding when this was determined by antagonism of Pho-A-dependent inhibition. These properties suggest the catalytic unit has both polar head group and acyl chain requirements for phospholipid binding.
Krall
J F
JF
UCLA-SFVP Department of Medicine, Veterans Administration Medical Center, Sepulveda 91343.
Leshon
S C
SC
Korenman
S G
SG
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D005260:Female; D006150:Guanine Nucleotides / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D009130:Muscle, Smooth / Q000201:enzymology*; D010743:Phospholipids / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism
D006150:Guanine Nucleotides; D010743:Phospholipids; D006165:Guanylyl Imidodiphosphate; D008345:Manganese; D000262:Adenylyl Cyclases; D008274:Magnesium