3d7m
Crystal Structure of the G Protein Fast-Exchange Double Mutant I56C/Q333CCrystal Structure of the G Protein Fast-Exchange Double Mutant I56C/Q333C
Structural highlights
FunctionGNAI1_RAT Guanine nucleotide-binding proteins (G proteins) are involved as modulators or transducers in various transmembrane signaling systems. The G(i) proteins are involved in hormonal regulation of adenylate cyclase: they inhibit the cyclase in response to beta-adrenergic stimuli. The inactive GDP-bound form prevents the association of RGS14 with centrosomes and is required for the translocation of RGS14 from the cytoplasm to the plasma membrane. May play a role in cell division.[1] Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedABSTRACT. Heterotrimeric G proteins (Galphabetagamma) transmit signals from activated G protein coupled receptors (GPCRs) to downstream effectors through a guanine nucleotide signaling cycle. Numerous studies indicate that the carboxy-terminal alpha5 helix of Galpha subunits participate in Galpha-receptor binding, and previous EPR studies suggest this receptor-mediated interaction induces a rotation and translation of the alpha5 helix of the Galpha subunit [Oldham et al., Nat. Struct. Mol. Biol., 13: 772-7 (2006)]. Based on this result, an engineered disulfide bond was designed to constrain the alpha5 helix of Galphai1 into its EPR-measured receptor-associated conformation through the introduction of cysteines at positions 56 in the alpha1 helix and 333 in the alpha5 helix (I56C/Q333C Galphai1). A functional mimetic of the EPR-measured alpha5 helix dipole movement upon receptor association was additionally created by introduction of a positive charge at the amino-terminus of this helix, D328R Galphai1. Both proteins exhibit dramatically elevated basal nucleotide exchange. The 2.9 A resolution crystal structure of the I56C/Q333C Galphai1 in complex with GDP-AlF4- reveals the shift of the alpha5 helix toward the guanine nucleotide-binding site that is anticipated by EPR measurements. The structure of the I56C/Q333C Galphai1 subunit further revealed altered positions for the switch regions and throughout the Galphai1 subunit, accompanied by significantly elevated crystallographic temperature factors. Combined with previous evidence in the literature, the structural analysis supports the critical role of electrostatics of the alpha5 helix dipole and overall conformational variability during nucleotide release. Helix dipole movement and conformational variability contribute to allosteric GDP release in Gi subunits.,Preininger A, Funk M, Meier S, Oldham W, Johnston C, Adhikary S, Kimple A, Siderovski D, Hamm H, Iverson T Biochemistry. 2009 Feb 17. PMID:19222191[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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