Dataset Information

Ric-8A, a GEF, and a Chaperone for G Protein ?-Subunits: Evidence for the Two-Faced Interface.

ABSTRACT: Resistance to inhibitors of cholinesterase 8A (Ric-8A) is a prominent non-receptor GEF and a chaperone of G protein ?-subunits (G?). Recent studies shed light on the structure of Ric-8A, providing insights into the mechanisms underlying its interaction with G?. Ric-8A is composed of a core armadillo-like domain and a flexible C-terminal tail. Interaction of a conserved concave surface of its core domain with the G? C-terminus appears to mediate formation of the initial Ric-8A/G?GDP intermediate, followed by the formation of a stable nucleotide-free complex. The latter event involves a large-scale dislocation of the G? ?5-helix that produces an extensive primary interface and disrupts the nucleotide-binding site of G?. The distal portion of the C-terminal tail of Ric-8A forms a smaller secondary interface, which ostensibly binds the switch II region of G?, facilitating binding of GTP. The two-site G? interface of Ric-8A is distinct from that of GPCRs, and might have evolved to support the chaperone function of Ric-8A.

SUBMITTER: Srivastava D

PROVIDER: S-EPMC7034654 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Publications

Ric-8A, a GEF, and a Chaperone for G Protein α-Subunits: Evidence for the Two-Faced Interface.

Srivastava Dhiraj D Artemyev Nikolai O NO

BioEssays : news and reviews in molecular, cellular and developmental biology 20200122 3

Resistance to inhibitors of cholinesterase 8A (Ric-8A) is a prominent non-receptor GEF and a chaperone of G protein α-subunits (Gα). Recent studies shed light on the structure of Ric-8A, providing insights into the mechanisms underlying its interaction with Gα. Ric-8A is composed of a core armadillo-like domain and a flexible C-terminal tail. Interaction of a conserved concave surface of its core domain with the Gα C-terminus appears to mediate formation of the initial Ric-8A/GαGDP intermediate, ...[more]

PMID: 31967346

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Project description:Resistance to Inhibitors of Cholinesterase A (Ric-8A) is a 60-kDa cytosolic protein that has chaperone and guanine nucleotide exchange (GEF) activity toward heterotrimeric G protein α subunits of the i, q, and 12/13 classes, catalyzing the release of GDP from Gα and subsequent binding of GTP. In the absence of GTP or GTP analogs, and subsequent to GDP release, Gα forms a stable nucleotide-free complex with Ric-8A. In this study, time-resolved fluorescence anisotropy measurements were employed to detect local motions of Gαi1 labeled at selected sites with Alexa 488 (C5) fluorescent dye (Ax) in the GDP, GTPγS (collectively, GXP), and Ric-8A-bound states. Sites selected for Alexa 488 (C5) derivatization were in the α-helical domain (residue 106), the α-helical domain-Ras-like domain hinge (residue 63), Switch I (residue 180), Switch II (residue 209), Switch III (residue 238), the α4 helix (residue 305), and at the junction between the purine-binding subsite in the β6-α5 loop and the C-terminal α helix (residue 330). In the GXP-bound states, the Alexa fluorophore reports local motions with correlation times ranging from 1.0 to 1.8 ns. The dynamics at Ax180 is slower in Gαi1•GDP than in Gαi1•GTPγS. The reverse is true at Ax209. The order parameters, S(2), for Alexa probes at switch residues are high (0.78-0.88) in Gαi1•GDP and lower (0.67-0.75) in Gαi1•GTPγS, although in crystal structures, switch segments are more ordered in the latter. Local motions at Ax63, Ax180, Ax209, and Ax330 are all markedly slower (2.3-2.8 ns) in Gαi1:Ric-8A than in Gαi1•GXP, and only modest (± 0.1) differences in S(2) are observed at most sites in Gαi1:Ric-8A relative to Gαi1•GXP. The slow dynamics suggests long-range correlated transitions within an ensemble of states and, particularly in the hinge and switch segments that make direct contact with Ric-8A. Induction of Gαi1 structural heterogeneity by Ric-8A provides a mechanism for nucleotide release.

Dataset Information

Ric-8A, a GEF, and a Chaperone for G Protein ?-Subunits: Evidence for the Two-Faced Interface.

Publications

Ric-8A, a GEF, and a Chaperone for G Protein α-Subunits: Evidence for the Two-Faced Interface.

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