A structural role for Asp83 in the photoactivation of rhodopsin.
Asp83 is a highly conserved residue in the second transmembrane domain of visual pigments and many members of other G protein-coupled receptor subfamilies. Upon illumination, the rod visual pigment rhodopsin proceeds through various intermediate states (Batho<-->BSI<-->Lumi<-->Meta I<-->Meta II). Meta II represents the active state of rhodopsin, which binds and activates the G protein transducin. Evidence has been presented that Asp83 participates in the formation of Meta II and undergoes a change in H-bonding. To investigate whether this role of Asp83 requires its proton-donating capacity and/or its H-bonding capability, we constructed the mutants D83C and D83N. Both mutants appear to effectively activate transducin, indicating that Asp83 is not essential for signal transduction. Differential effects of the mutations D83C and D83N are observed in the spectral properties and the pH sensitivity of the Meta I-->Meta II transition. In general, D83C behaves much more like wild-type than D83N. We conclude that the structural role of Asp83 also involves the acidic nature of its carboxyl group. In addition, the participation in Meta II formation of Cys83 in D83C manifests itself as a change in the vibrational properties of the sulfhydryl group, demonstrating that the -SH group can be used as a non-invasive probe for local structural changes.
Breikers
G
G
Department of Biochemistry, Nijmegen Centre for Molecular Life Sciences, University of Nijmegen, The Netherlands.
Bovee-Geurts
P H
PH
DeCaluwé
G L
GL
DeGrip
W J
WJ
D000595:Amino Acid Sequence; D001224:Aspartic Acid; D019204:GTP-Binding Proteins / Q000378:metabolism; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D010777:Photochemistry; D017354:Point Mutation; D011487:Protein Conformation; D017510:Protein Folding; D012243:Rhodopsin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017550:Spectroscopy, Fourier Transform Infrared
D001224:Aspartic Acid; D012243:Rhodopsin; D019204:GTP-Binding Proteins