Format

Send to

Choose Destination
Biochemistry. 2004 Sep 7;43(35):11153-62.

Probing rhodopsin-transducin interactions by surface modification and mass spectrometry.

Author information

1
Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, South Carolina 29425, USA.

Abstract

The interactions of rhodopsin and the alpha-subunit of transducin (G(t)) have been mapped using a surface modification "footprinting" approach in conjunction with mass spectrometric analysis employing a synthetic peptide corresponding to C-terminal residues 340-350 of the alpha-subunit of G(t), G(t)alpha(340-350). Membrane preparations of unactivated (Rh) and light-activated rhodopsin (Rh*), each in the presence or absence of G(t)alpha(340-350), were acetylated with the water-soluble reagent sulfosuccinimidyl acetate, and the extent of the acetylation was determined by mass spectrometry. By comparing the differences in acetylation among Rh, Rh*, and the Rh-G(t)alpha(340-350) and Rh*-G(t)alpha(340-350) complexes, we demonstrate that the surface exposure of the acetylation sites was reduced by the conformational change associated with light activation, and that binding of G(t)alpha(340-350) blocks acetylation sites on cytoplasmic loops 1, 2, and 4 of Rh*. In addition, we show evidence of interaction between the end of the C-terminal tail of rhodopsin and G(t)alpha in the unactivated state of rhodopsin.

PMID:
15366925
DOI:
10.1021/bi049642f
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for American Chemical Society
Loading ...
Support Center