We have used surface plasmon resonance (SPR) measurements for the kinetic analysis of G-protein-receptor interaction monitored in real time. Functionally active rhodopsin was immobilized on an SPR surface, with full retention of biochemical specific activity for catalysis of nucleotide exchange on the retinal G-protein alpha subunit, via binding to immobilized concanavalin A. The binding intera...

Rhodopsin is the only G protein-coupled receptor (GPCR) whose 3D structure is known; therefore, it serves as a prototype for studies of the GPCR family of proteins. Rhodopsin dysfunction has been linked to misfolding, caused by chemical modifications that affect the naturally occurring disulfide bond between C110 and C187. Here, we identify the structural elements that stabilize rhodopsin by co...

G protein-coupled receptors (GPCRs) are seven transmembrane domain proteins that transduce extracellular signals across the plasma membrane and couple to the heterotrimeric family of G proteins. Like most intrinsic membrane proteins, GPCRs are capable of oligomerization, the function of which has only been established for a few different receptor systems. One challenge in understanding the func...

We have conducted a comprehensive analysis of the relative timing of the terminal mitosis and the onset of rhodopsin expression in rod precursors in the rat retina in vivo. This analysis demonstrated that there are two distinct phases of rod development during retinal histogenesis. For the majority of rod precursors, those born on or after embryonic day 19 (E19), the onset of rhodopsin expressi...

Rhodopsin has been used as a prototype system to investigate G protein-coupled receptor (GPCR) internalization and endocytic sorting mechanisms. Failure of rhodopsin recycling upon light activation results in various degenerative retinal diseases. Accumulation of internalized rhodopsin in late endosomes and the impairment of its lysosomal degradation are associated with unregulated cell death t...

The lectin chaperone calnexin (Cnx) is important for quality control of glycoproteins, and the chances of correct folding of a protein increase the longer the protein interacts with Cnx. Mutations in glycoproteins increase their association with Cnx, and these mutant proteins are retained in the endoplasmic reticulum. However, until now, the increased interaction with Cnx was not known to incre...

A variety of techniques are currently in use for preparing protein-containing lipid vesicles known as proteoliposomes. However, the functionality of membrane protein in proteoliposomes prepared by various techniques has rarely been evaluated directly. We prepared rhodopsin-containing proteoliposomes consisting of asolectin or native retinal rod outer segment disk lipids using n-octyl beta-d-glu...

MOTIVATION Rhodopsin is a visual pigment present in rod cells of retina. It belongs to GPCR family and involves photoisomerization of 11-cis-retinal to all-trans-retinal isomers, conformational changes in rhodopsin and signal transduction cascade to generate a nerve impulse. This signaling pathway has been targeted to eliminate the effect of a mutation (Gly90→Asp) responsible for abnormal activ...

In vertebrate retinal rods, rhodopsin undergoes a conformational change upon absorption of a quantum of light. The primary photoprocess is followed by a transient depolarisation of the outer receptor membrane. It is generally accepted that the depolarisation is due to the closing of Na’channels [l-4]. As the rhodopsin containing inner discs and the plasma membrane of the retinal rods seem to be...

PURPOSE To test the hypothesis that a lack of Tubby-like protein 1 (TULP1) function causes aberrant transport of nascent rhodopsin and to examine the functional relationship between the homologous proteins TULP1 and Tubby by studying mice carrying combined mutations. METHODS Subcellular localization of TULP1 and rhodopsin in photoreceptors was determined by immunofluorescence and by postembed...