Calcium/calmodulin-regulated guanylate cyclase of the excitable ciliary membrane from Paramecium. Dissociation of calmodulin by La3+: calmodulin specificity and properties of the reconstituted guanylate cyclase.
Ca2+-regulated guanylate cyclase in ciliary membranes from Paramecium contained tightly bound calmodulin. Antisera against calmodulin from Tetrahymena and soybean inhibited enzyme activity. EGTA did not easily release calmodulin; however, La3+ inhibited guanylate cyclase by dissociation of calmodulin. While La could not replace Ca in the activation of guanylate cyclase, it substituted for Ca2+ in the activation of calmodulin-dependent phosphodiesterase from pig brain independently of whether homologous or Paramecium calmodulin was used. After removal of endogenous calmodulin from guanylate cyclase, reconstitution was achieved with calmodulin from Paramecium, Tetrahymena, pig brain, and soybean. Ca2+-binding proteins lacking trimethyllysine like calmodulin from Dictyostelium, parvalbumin, and troponin C failed to restore enzyme activity. The properties of the native and reconstituted guanylate cyclase/calmodulin complex were compared. Reassociation of calmodulin with its target enzyme was weak since all calmodulin remained in the supernatant after a single centrifugation. While most enzyme characteristics remained unchanged in the reconstituted complex, the inhibition by Ca greater than 100 microM was of a mixed-type compared to noncompetitive inhibition in the native enzyme. The regulation of the enzyme by cations was also altered. Whereas Ca was the most potent and specific activator of the native enzyme, in the reconstituted system Sr was far more effective.
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000494:pharmacology* / Q000502:physiology; D002413:Cations, Divalent; D002462:Cell Membrane / Q000201:enzymology; D002923:Cilia / Q000201:enzymology*; D006162:Guanylate Cyclase / Q000378:metabolism*; D007700:Kinetics; D007811:Lanthanum / Q000494:pharmacology*; D010247:Paramecium / Q000201:enzymology*; D013045:Species Specificity
D002147:Calmodulin; D002413:Cations, Divalent; D007811:Lanthanum; D006162:Guanylate Cyclase; D002118:Calcium