Covalent and noncovalent interactions mediate metabotropic glutamate receptor mGlu(5) dimerization

Some, perhaps all, G protein-coupled receptors form homo- or heterodimers. We have shown that metabotropic glutamate receptors are covalent dimers, he ld together by one or more disulfide bonds near the N terminus. Here we rep ort how mutating cysteines in this region affect dimerization and function. Covalent dimerization is preserved when cysteines 57, 93, or 99 are mutate d but lost with replacement at 129. Coimmunoprecipitation under nondenaturi ng conditions indicates that the C[129]S mutant receptor remains a dimer, v ia noncovalent interactions. Both C[93]S and C[129]S bind [H-3] quisqualate , whereas binding to C[57]S or C[99]S mutants is absent or greatly attenuat ed. The C[93]S and C[129]S receptors have activity similar to wild-type whe n assayed by fura-2 imaging of intracellular calcium in human embryonic kid ney cells or electrophysiologically in Xenopus laevis oocytes. In contrast, C[57]S or C[99]S are less active in both assays but do respond with higher glutamate concentrations in the oocyte assay. These results demonstrate th at 1) covalent dimerization is not critical for mGlu(5) binding or function ; 2) mGlu(5) remains a noncovalent dimer even in the absence of covalent di merization; and 3) high-affinity binding requires Cys-57 and Cys-99.