Structures of the intradiskal loops and amino terminus of the G‐protein receptor, rhodopsin

Abstract

The intradiskal surface of the transmembrane protein, rhodopsin, consists of the amino terminal domain and three loops connecting six of the seven transmembrane helices. This surface corresponds to the extracellular surface of other G-protein receptors. Peptides that represent each of the extramembraneous domains on this surface (three loops and the amino terminus) were synthesized. These peptides also included residues which, based on a hydrophobic plot, could be expected to be part of the transmembrane helix. The structure of each of these peptides in solution was then determined using two-dimensional 1H nuclear magnetic resonance. All peptide domains showed ordered structures in solution. The structures of each of the peptides from intradiskal loops of rhodopsin exhibited a turn in the central region of the peptide. The ends of the peptides show an unwinding of the transmembrane helices to form this turn. The amino terminal domain peptide exhibited alpha-helical regions with breaks and bends at proline residues. This region forms a compact domain. Together, the structures for the loop and amino terminus domains indicate that the intradiskal surface of rhodopsin is ordered. These data further suggest a structural motif for short loops in transmembrane proteins. The ordered structures of these loops, in the absence of the transmembrane helices, indicate that the primary sequences of these loops are sufficient to code for the turn.

Biosynthesis and NMR-studies of a double transmembrane domain from the Y4 receptor, a human GPCR

“…A large body of literature supports the basic assumption of the model: For example, proteolysis of membrane proteins resulted in fragments containing entire TM sequences (Huang et al 1981), and chemically or recombinantly synthesized TM peptides spontaneously assembled thereby rescuing receptor activity (Kahn and Engelman 1992;Ridge et al 1995;Martin et al 1999;Wrubel et al 1994). Finally, peptides corresponding to the N and C terminus (Harmar 2001;O'Hara et al 1993), loop domains (Bennett et al 2004;Katragadda et al 2001a, b;Yeagle et al 2000) and transmembrane domains (Katragadda et al 2001a, b;Cohen et al 2008;Zheng et al 2006;Musial-Siwek et al 2008;Tian et al 2007;Lau et al 2008;Mobley et al 2007;Neumoin et al 2007) from GPCRs have been found to fold to distinct secondary structures which in certain cases resembled the structures of the corresponding regions of the intact receptor.…”

Specific inhibition of HIV-1 coreceptor activity by synthetic peptides corresponding to the predicted extracellular loops of CCR5

“…Kennedy and E.A.B., unpublished observations, July 2003) for a chimera containing the N-terminus of CCR5 grafted onto the corresponding region of bacteriorhodopsin, 41 a 7-transmembrane receptor with structural features dissimilar to those of mammalian GPCRs. The recent demonstration of ordered structures for soluble peptides representing each of the extracellular regions of bovine rhodopsin 42 suggests that considerable insight might be gained from structural analysis of peptides derived from the extracellular regions of CCR5 and from comparisons with the structures of peptides from the corresponding regions of CXCR4 and other GPCRs, with and without coreceptor activity. The synthetic peptide approach thus has potential to bring order to the bewildering complexity of HIV-1 Env/coreceptor interactions.…”

Electrostatic considerations affecting the calculated HOMO–LUMO gap in protein molecules

“…Turning our attention to polypeptide systems, six protein conformations (1PLW [45], 1FUL [46], 1RVS [47], 1EDW [48], 1FDF [49] and 1UBQ [50]) from the Brookhaven National Laboratory Protein Data Bank (PDB) [51] were used as starting configurations for our calculations. Table 1 shows the computed HOMO-LUMO gaps for these structures in vacuo, using the PBE functional.…”