Structural and Functional Role of Helices I and II in Rhodopsin

Abstract

The naturally occurring mutations G51A and G51V in transmembrane helix I and G89D in the transmembrane helix II of rhodopsin are associated with the retinal degenerative disease autosomal dominant retinitis pigmentosa. To probe the orientation and packing of helices I and II a number of replacements at positions 51 and 89 were prepared by using site-directed mutagenesis, and the corresponding proteins expressed in COS-1 cells were characterized. Mutations at position 51 (G51V and G51L) bound retinal like wild-type rhodopsin but had thermally destabilized structures in the dark, altered photobleaching behavior, destabilized metarhodopsin II active conformations, and were severely defective in signal transduction. The effects observed can be correlated with the size of the mutated side chains that would interfere with specific interhelical interaction with Val-300 in helix VII. Mutations at position 89 had sensitivity to charge, as in G89K and G89D mutants, which showed reduced transducin activation. G89K showed a second absorbing species in the UV region at 350 nm, suggesting a charge effect of the introduced lysine. Increased formation of non-active forms of rhodopsin, like metarhodopsin III, may have some influence in the molecular defect underlying retinitis pigmentosa in the mutants studied. At the structural level, the effect of the mutations analyzed can be rationalized assuming a very specific set of tertiary interactions in the interhelical packing of the transmembrane segments of rhodopsin.

Coupling of Human Rhodopsin to a Yeast Signaling Pathway Enables Characterization of Mutations Associated with Retinal Disease

“…As M39R rhodopsin is expressed more productively by mammalian cells than P23H rhodopsin, and a proportion remains able to form a light-responsive complex with retinal, it was selected as an intermediate RP-associated rhodopsin mutation (Davies et al 2012;Ramon et al 2014). G51A is the most common nonsynonymous rhodopsin mutation in humans (Lek et al 2016), displays a less severe phenotype in vitro and in patients (Cideciyan et al 1998;Bosch et al 2003), and may be an asymptomatic variant (Athanasiou et al 2018). These three rhodopsin mutants were each expressed using the mCherry reporter yeast strain.…”

Transmembrane helix: simple or complex

“…To recapitulate, the functional role of TM-5 in the latter has not been established whereas the Gly51 in TM-1 and Gly89 in TM-2 have been linked to the retinal degenerative disease autosomal dominant retinitis pigmentosa (35) while Glu113 in TM-3, Ala169 in TM-4, Trp265 in TM-6 and Lys296 in TM7 are functionally important (36,37). Indeed, only TM-5 [positions 200–225 (38); z -score of −6.12] in bovine rhodopsin is considered simple by TMSOC and was masked in the sequence.…”

A Novel Rhodopsin Gene from Octopus vulgaris for Optobioelectronics Materials

“…The homology model of melanopsin (maximal absorbance λmax = 447 nm) based on the crystal structure of squid rhodopsin (λmax = 490 nm) shows that 43 nm spectral shift is due to increased bond-length alternation of the protonated Schiff base of 11-cis-retinal chromophore, induced by N87Q mutation and water-mediated H-bonding interactions with the Schiff base linkage [19]. This phenomenon is analogous to spectral changes observed in the G89Q bovine rhodopsin mutants [32]. The alignment with novel OctR structure reveals N88 position for mutagenesis for developing "blue light" bistable recombinant Rho.…”