Dimerization of guanylyl cyclase-activating protein and a mechanism of photoreceptor guanylyl cyclase activation

Abstract

Ca2+-binding guanylyl cyclase-activating proteins (GCAPs) stimulate photoreceptor membrane guanylyl cyclase (retGC) in the light when the free Ca2+ concentrations in photoreceptors decrease from 600 to 50 nM. RetGC activated by GCAPs exhibits tight dimerization revealed by chemical cross- linking (Yu, H., Olshevskaya, E., Duda, T., Seno, K., Hayashi, F., Sharma, R. K., Dizhoor, A.M., and Yamazaki, A. (1999) J. Biol. Chem. 274, 15547-15555). We have found that the Ca2+-loaded GCAP-2 monomer undergoes reversible dimerization upon dissociation of Ca2+. The ability of GCAP-2 and its several mutants to activate retGC in vitro correlates with their ability to dimerize at low free Ca2+ concentrations. A constitutively active GCAP-2 mutant E80Q/E116Q/D158N that stimulates retGC regardless of the free Ca2+ concentrations forms dimers both in the absence and in the presence of Ca2+. Several GCAP-2/neurocalcin chimera proteins that cannot efficiently activate retGC in low Ca2+ concentrations are also unable to dimerize in the absence of Ca2+. Additional mutation that restores normal activity of the GCAP-2 chimera mutant also restores its ability to dimerize in the absence of Ca2+. These results suggest that dimerization of GCAP-2 can be a part of the mechanism by which GCAP-2 regulates the photoreceptor guanylyl cyclase. The Ca2+-free GCAP-1 is also capable of dimerization in the absence of Ca2+, but unlike GCAP-2, dimerization of GCAP-1 is resistant to the presence of Ca2+.