Structure and Function Study of Melanopsin

Christine Shieh
University of Maryland, Baltimore County

Melanopsin is an unusual opsin-like protein expressed in a subset of retinal ganglion cells (RGCs). The axonal projections of these RGCs form the retinohypothalamic tract, which sends input from the retina to the suprachiasmatic nuclei (SCN). The SCN is known to be integral to light entrainment of the mammalian circadian clock.

Melanopsin is believed to be the elusive photopigment that is involved in regulating circadian rhythms and photoentrainment. However, melanopsin absorbs at a wavelength of 420 nm in vitro, and the photopigment mediating the electrical response in the subset of RGCs absorbs at 479 nm. The spectral properties do not match. Subsequently, it is necessary to investigate the photochemical, spectral and signaling properties of melanopsin to see if they can be used to elucidate its role in the SCN-projecting RGCs.

In addition, although mouse melanopsin is a vertebrate photopigment, it is more closely related to cephalopod rhodopsin (~39% identical) and other inverterbrate opsins than it is to vertebrate rod and cone opsins (~ 30%). It is most likely that melanopsin’s unique structural motifs underlie its unusual photochemical properties and it will share some of these important motifs with invertebrate rhodopsins.

My goal is to create mutations in melanopsin in order to examine how changes in the binding pocket residues affect the absorbance spectrum of melanopsin and its activated intermediate. In addition, I will create corresponding mutations in rhodopsin in order to understand its impact on meta-rhodopsin II, the activated rhodopsin state. Using site-directed mutagenesis, I will create a mutation at the amino acid site 122 in melanopsin and rhodopsin. In melanopsin there is an isoleucine at site 122, similar to invertebrate rhodopsin. In rhodopsin, there is a glutamate. Glutamate interacts with the ß-ionone ring of the chromophore 11-cis retinal and controls the lifetime of Meta-Rhodopsin II. Replacing the charged amino acid at site 122 could influence the stability of the active photopigment and the absorbance of the photopigment .