| Abstract
| - The evolutionary trace (ET) method, a data mining approach for determining significant levelsof amino acid conservation, has been applied to over 700 aligned G-protein-coupled receptor(GPCR) sequences. The method predicted the occurrence of functionally important clusters ofresidues on the external faces of helices 5 and 6 for each family or subfamily of receptors;similar clusters were observed on helices 2 and 3. The probability that these clusters are notrandom was determined using Monte Carlo techniques. The cluster on helices 5 and 6 isconsistent with both 5,6-contact and 5,6-domain swapped dimer formation; the possibleequivalence of these two types of dimer is discussed because this relates to activation by homo-and heterodimers. The observation of a functionally important cluster of residues on helices 2and 3 is novel, and some possible interpretations are given, including heterodimerization andoligomerization. The application of the evolutionary trace method to 113 aligned G-proteinsequences resulted in the identification of two functional sites. One large, well-defined site isclearly identified with adenyl cyclase, β/γ and regulator of G-protein signaling (RGS) binding.The other G-protein functional site, which extends from the ras-like domain onto the helicaldomain, has the correct size and electrostatic properties for GPCR dimer binding. Theimplications of these results are discussed in terms of the conformational changes required inthe G-protein for activation by a receptor dimer. Further, the implications of GPCR dimerizationfor medicinal chemistry are discussed in the context of these ET results.
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