# Mechanism of G protein Activation by Ric-8A

> **NIH NIH R01** · UNIVERSITY OF MONTANA · 2020 · $362,500

## Abstract

PROJECT SUMMARY R01GM105993
The goal of this renewal of R01-GM105993 is to determine the structure and dynamic properties of the
complex between Ric-8A and the alpha subunit of the heterotrimeric G protein Gi (Gαi1). Heterotrimeric G
proteins modulate cell metabolism, secretion, electrical conductivity, gene transcription, cell division and
cellular motility, and therefore are essential to eukaryotic life. Misregulation of G proteins is associated with
cancer and a range of other diseases of relevance to general medicine. While most processes controlled by
heterotrimeric G proteins occur at cell membranes, recent research has shown that G alpha subunits (Gα) also
control certain events in the cell cytoplasm. Important among these is asymmetric cell division, which is
essential for embryonic development. Ric-8A is critical regulator of Gα in this process. Ric-8A is a Guanine
nucleotide Exchange Factor (GEF) that activates Gα by catalyzing the exchange of guanosine diphosphate
(GDP) for guanosine triphosphate (GTP) at the active site of Gα. The intermediate in this reaction is the
nucleotide-free Gα:Ric-8A complex. Describing the structural changes that occur when Ric-8A binds to
Gα·GDP is key to understanding how Ric-8A activates Gα. Ric-8A is also a chaperone that promotes proper
folding of Gα in cells. The two aims of this proposal will test the hypothesis that Gαi1 and possibly Ric-8A
sample multiple conformational states in the complex Gαi1:Ric-8A and to make use of structural information to
ask whether the GEF and chaperone activities are mechanistically inseparable, or are distinct functions that
can be decoupled. The mechanism by which Ric-8A is regulated by protein kinase CK2 will also be
investigated. Aim 1 is to determine the structure of Ric-8A and its complex with Gαi1 by X-ray crystallography,
with the former in both phosphorylated and non-phosphorylated states. Camelid single chain heavy chain
variable domains will be used as crystallization adjuvants. Collaborative heteronuclear NMR experiments will
be conducted as an independent approach to define the interface between Ric-8A and GαI and to identify
residues in transition among conformational states. Assays of GEF and chaperone activities will be conducted
to assess the role of residues that are hypothesized from structural studies as mediators of one or both of
these activities. Aim 2 will determine the global structure of Ric-8A:Gαi1 using small angle X-ray scattering
and cryo-electron microscopy. Modeling of the lower resolution structures using molecular dynamics and
Monte Carlo simulations with X-ray structures of Ric-8A and Gαi1 will be used to detect and identify multiple
conformational states of the complex.

## Key facts

- **NIH application ID:** 9989171
- **Project number:** 5R01GM105993-08
- **Recipient organization:** UNIVERSITY OF MONTANA
- **Principal Investigator:** Stephen R Sprang
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $362,500
- **Award type:** 5
- **Project period:** 2013-04-01 → 2023-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9989171

## Citation

> US National Institutes of Health, RePORTER application 9989171, Mechanism of G protein Activation by Ric-8A (5R01GM105993-08). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9989171. Licensed CC0.

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