Structure and function in rhodopsin: Further elucidation of the role of the intradiscal cysteines, Cys-110, -185, and -187, in rhodopsin folding and function

The disulfide bond between Cys-110 and Cys-187 in the intradiscal domain is required for correct folding in vivo and function of mammalian rhodopsin. Mis-folding in rhodopsin, characterized by the loss of ability to bind 11-c is-retinal, has been shown to be caused by an intradiscal disulfide bond di fferent from the above native disulfide bond. Further, naturally occurring single mutations of the intradiscal cysteines (C110F, C110Y and C187Y) are associated with retinitis pigmentosa (RP). To elucidate further the role of every one of the three intradiscal cysteines, mutants containing single-cy steine replacements by alanine residues and the above three RP mutants have been studied. We find that C110A, C110F, and C1110Y all form a disulfide b ond between C185 and C187 and cause loss of retinal binding. C185A allows t he formation of a C110-C187 disulfide bond, with wild-type-like rhodopsin p henotype. C187A forms a disulfide bond between C110 and C185 and binds reti nal, and the pigment formed has markedly altered bleaching behavior. Howeve r, the opsin from the RP mutant C187Y forms no rhodopsin chromophore.