The quest to understand heterotrimeric G protein signaling

Abstract

Over the past fifteen years, great strides have been made to understand how heterotrimeric G proteins control their downstream targets. However, the mechanism by which heterotrimeric G proteins are activated by their upstream G protein-coupled receptors (GPCRs) remains obscure. Recent structural data support the idea that GPCRs, despite their small size, are sophisticated allosteric machines with multiple signaling outputs.

Figure 1

The heterotrimeric G protein cycle. The inactive Gαβγ heterotrimer (top) is composed of two principal elements, Gα·GDP (cyan, with GDP shown in grey with magenta dot surface) and the Gβγ heterodimer (blue and green). Gβγ sequesters the switch II element (red) such that it is unable to interact with effectors. Activated GPCRs catalyze the release of GDP from Gα, allowing GTP to bind and liberate the activated Gα·GTP subunit (right). In this state, switch II forms a helix stabilized by the γ-phosphate of GTP. The activated Gαsubunit can then interact with effectors, such as the catalytic domains of adenylyl cyclase (bottom, gold and dark green). Switch II forms a major component of the interface, as it does in other characterized effector complexes. The Gαsubunit has a slow GTPase activity that converts GTP to GDP, weakening its interactions with effectors and allowing it to dissociate as a deactivated Gα·GDP subunit (left). In this state, switch II is disordered (red dotted line), and the protein has high affinity for Gβγ subunits, completing the cycle. The structures shown correspond to PDB entries 1GG2 (ref. ²⁷) (top, cyan subunit corresponds to Gα_i), 1AZT²⁸ (right, Gα_s), 1AZS⁴ (bottom, Gα_s) and 1GDD²⁹ (left, Gα_i).

Figure 2

Structural transitions of GPCRs and their interactions with heterotrimeric G proteins. (a) Comparison of the structures of rhodopsin and opsin. Opsin represents a retinal-depleted form of the GPCR with low basal activity yet shows structural features consistent with changes predicted to occur during rhodopsin activation, such as the outward twist of the third intracellular loop (IL3). Opsin can also be crystallized in complex with a peptide derived from the C-terminus of Gα_t (yellow), the region of the heterotrimer most strongly linked to receptor recognition. Retinal is shown as a stick model with a green dot surface. Structures correspond to PDB entries 1GZM³⁰ and 3DQB¹⁷. (b) Conceptual problems in docking current models of GPCRs with their heterotrimeric G protein substrates. Here we show the inactive Gα_iβ₁γ₂ complex (PDB 1GG2 (ref. ²⁷)), in which the C-terminus of Gα_i was extended according to PDB entry 1AGR³¹ and the C-terminus of Gγ according to PDB entry 1OMW³². The C-terminal span of Gα_i analogous to the Gα_t peptide bound to opsin is colored yellow. Comparing the orientation of this helix in each model reveals an an apparent docking incompatibility, because the intact heterotrimer would have to be rotated, roughly counterclockwise, up into the plane of the lipid bilayer in order to superimpose these elements. (c) Collision of Gαβγ with the lipid bilayer when superimposed with the Gα_t peptide bound to opsin. The collision suggests that either the model of opsin does not represent a GPCR in a fully activated state, or the G protein heterotrimer must undergo a significant conformational change, or both. A large conformational change is expected in Gαβγ because its interaction with receptors must induce nucleotide release.