CALCIUM-BINDING, BUT NOT A CALCIUM-MYRISTOYL SWITCH, CONTROLS THE ABILITY OF GUANYLYL CYCLASE-ACTIVATING PROTEIN GCAP-2 TO REGULATE PHOTORECEPTOR GUANYLYL CYCLASE

Guanylyl cyclase-activating protein 2 (GCAP-2) is a recoverin-like cal cium binding protein that regulates photoreceptor guanylyl cyclase (Re tGC) (Dizhoor, A, M., and Hurley, J. B. (1996) J. Biol. Chem. 271, 193 46-19350), It was reported that myristoylation of a related protein, G CAP-1, was critical for its affinity for RetGC (Frins, S,, Bonigk W,, Muller, F,, Kellner, R,, and Koch, K,-W, (1996) J, Biol. Chem, 271, 80 22-8027), We demonstrate that the N terminus of GCAP-2, like those of other members of the recoverin family of Ca2+-binding proteins, is fat ty acylated. However, unlike other proteins of this family, more GCAP- 2 is present in the membrane fraction at low Ca2+ than at high Ca2+ co ncentrations. We investigated the role of the N-terminal fatty acyl re sidue in the ability of GCAP-2 to regulate RetGCs, Myristoylated or no nacylated GCAP-2 forms were expressed in Escherichia coli, Wild-type G CAP-2 and the Gly(2) --> Ala(2) GCAP-2 mutant, which is unable to unde rgo N-terminal myristoylation, were also expressed in mammalian HEK293 cells, We found that compartmentalization of GCAP-2 in photoreceptor outer segment membranes is Ca2+- and ionic strength-sensitive, but it does not require the presence of the fatty acyl group and does not nec essarily directly reflect GCAP-2 interaction with RetGC, The lack of m yristoylation does not significantly affect the ability of GCAP-2 to s timulate RetGC, Nor does it affect the ability of the Ca2+-loaded form of GCAP-2 to compete with the GCAP-2 mutant that constitutively activ ates RetGC, We conclude that while Ca2+ binding plays a major regulato ry role in GCAP-2 function, it does not operate through a calcium-myri stoyl switch similar to the one found in recoverin.