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Updated: Jun 12, 2025
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Comparing the Affinity of GTPase-binding Proteins using Competition Assays
Published on: October 8, 2015
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A molten globule ensemble primes Arf1-GDP for the nucleotide switch
Tejaswi Koduru1, Noam Hantman2, Edgar V Peters3
1Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180.
Summary
Small GTPases like Arf1 switch between GDP and GTP states via conformational changes. Pressure induces a molten globule state, facilitating this nucleotide exchange, crucial for cell signaling.
Area of Science:
- Molecular biology
- Biophysics
- Cellular signaling
Background:
- Adenosine di-phosphate (ADP) ribosylation factor (Arf) GTPases act as molecular switches controlling cell membrane organization.
- The GDP/GTP nucleotide exchange in Arf1 requires external factors and involves significant conformational changes, but the mechanism remains unclear.
Purpose of the Study:
- To elucidate the mechanism of the energetically demanding GDP/GTP switch in Arf1.
- To investigate the role of conformational changes in Arf1 nucleotide exchange.
Main Methods:
- Utilized pressure perturbation combined with Nuclear Magnetic Resonance (NMR), Fourier Transform Infra-Red spectroscopy (FTIR), Small-Angle X-ray Scattering (SAXS), and fluorescence.
- Employed computational methods to analyze structural dynamics.
Main Results:
- Pressure induced the formation of a molten globule (MG) ensemble in Arf1.
- Pressure also promoted the GDP to GTP transition, indicating the MG ensemble's functional role in nucleotide switching.
Conclusions:
- The MG ensemble facilitates Arf1 nucleotide switching without complete unfolding, potentially serving as a recognition site for Guanine Nucleotide Exchange Factors (GEFs).
- This MG-based switching mechanism may be conserved across Arf GTPases, Arf-like GTPases, and related small GTPases like Rags and Gα GTPases.
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