Genetic and structural analysis of G protein α subunit regulatory domains

Abstract

Genetic and structural analysis of the alpha chain polypeptides of heterotrimeric G proteins defines functional domains for GTP/GDP binding, GTPase activity, effector activation, receptor contact and beta gamma subunit complex regulation. The conservation in sequence comprising the GDP/GTP binding and GTPase domains among G protein alpha subunits readily allows common mutations to be made for the design of mutant polypeptides that function as constitutive active or dominant negative alpha chains when expressed in different cell types. Organization of the effector activation, receptor and beta gamma contact domains is similar in the primary sequence of the different alpha subunit polypeptides relative to the GTP/GDP binding domain sequences. Mutation within common motifs of the different G protein alpha chain polypeptides have similar functional consequences. Thus, what has been learned with the Gs and Gi proteins and the regulation of adenylyl cyclase can be directly applied to the analysis of newly identified G proteins and their coupling to receptors and regulation of putative effector enzymes.

Activating mutations in the NH2‐ and COOH‐terminal moieties of the Gsα subunit have dominant phenotypes and distinguishable kinetics of adenylyl cyclase stimulation

“…Rather, the mutation of the COOH-terminus relieves an inhibition of the a, activation domain independent of the intrinsic GTPase activity of the (Y subunit polypeptide. The ability of COOHterminal mutations to influence the a, activation domain is not unexpected because the COOH-terminus is oriented towards the a subunit activation domain within the polypeptide tertiary structure [20,28] and is the major contact site for activated receptors [23], which function to enhance GDP dissociation allowing activation by GTP. The proximity of the COOHterminus to the a, activation domain could, therefore, influence tertiary conformations to alter the intrinsic properties of the a, polypeptide.…”

G-Proteins: implications for pathophysiology and disease

“…These observations invoke G-proteins as possible protooncogenes (3,4). G-Protein structure and properties G-Proteins are heterotrimers composed of three distinct subunits: a (39-46kD), ß (w37kD) and 7 («8kD) (2)(3)(4). The ß-and 7-subunits exist as a tightly associated complex (8) that represents a functional entity.…”

“…For instance, naturally occurring mutations in G-proteins have been detected in various endocrine tumours (4) and yield constitutively activated signalling systems (3,4). These observations invoke G-proteins as possible protooncogenes (3,4). G-Protein structure and properties G-Proteins are heterotrimers composed of three distinct subunits: a (39-46kD), ß (w37kD) and 7 («8kD) (2)(3)(4).…”

“…1) and abnormally G-protein-mediated transmembrane signalling may therefore play a key role in the precipitation of disease. For instance, naturally occurring mutations in G-proteins have been detected in various endocrine tumours (4) and yield constitutively activated signalling systems (3,4). These observations invoke G-proteins as possible protooncogenes (3,4).…”

“…Since then, the G-proteins have emerged as an expanding family of signal transducers, coupling diverse hormone receptors to intracellular effectors (2)(3)(4)(5). Quantitative and qualitative alterations in G-proteins may profoundly affect hormone action and lead to clinically manifest dysfunction of several organ systems.…”