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Crystal structure of the GTPase-activating domain of human p120GAP and implications for the interaction with Ras
K Scheffzek1, A Lautwein, W Kabsch
1Max-Planck-Institut für molekulare Physiologie, Dortmund, Germany.
Abstract:
Ras-related GTP-binding proteins function as molecular switches which cycle between GTP-bound 'on'- and GDP-bound 'off'-states. GTP hydrolysis is the common timing mechanism that mediates the return from the 'on' to the 'off'-state. It is usually slow but can be accelerated by orders of magnitude upon interaction with GTPase-activating proteins (GAPs). In the case of Ras, a major regulator of cellular growth, point mutations are found in approximately 30% of human tumours which render the protein unable to hydrolyse GTP, even in the presence of Ras-GAPs. The first structure determination of a GTPase-activating protein reveals the catalytically active fragment of the Ras-specific p120GAP (ref. 2), GAP-334, as an elongated, exclusively helical protein which appears to represent a novel protein fold. The molecule consists of two domains, one of which contains all the residues conserved among different GAPs for Ras. From the location of conserved residues around a shallow groove in the central domain we can identify the site of interaction with Ras x GTP. This leads to a model for the interaction between Ras and GAP that satisfies numerous biochemical and genetic data on this important regulatory process.
Insights
GTPase-activating proteins (GAPs) accelerate Ras protein GTP hydrolysis, regulating cell growth. Researchers determined the structure of a Ras-specific GAP fragment, revealing its interaction site with Ras-GTP and a novel protein fold.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Ras-related proteins are molecular switches regulating cellular processes.
- GTP hydrolysis, facilitated by GTPase-activating proteins (GAPs), is crucial for this switching mechanism.
- Mutations in Ras proteins are implicated in approximately 30% of human tumors, often impairing GTP hydrolysis.
Purpose of the Study:
- To determine the structure of the catalytically active fragment of the Ras-specific p120 GTPase-activating protein (GAP), designated GAP-334.
- To identify the interaction site between GAP-334 and Ras-GTP.
- To propose a model for Ras-GAP interaction based on structural and biochemical data.
Main Methods:
- X-ray crystallography for structure determination of GAP-334.
- Analysis of conserved residues within the GAP-334 structure.
- Integration of structural findings with existing biochemical and genetic data.
Main Results:
- GAP-334, the catalytically active fragment of p120GAP, adopts an elongated, exclusively helical structure, representing a novel protein fold.
- The protein comprises two domains, one containing residues conserved among Ras-specific GAPs.
- Conserved residues are located around a shallow groove, identified as the Ras-GTP interaction site.
Conclusions:
- The determined structure provides insights into the mechanism of Ras-GAP interaction.
- The findings elucidate how GAPs accelerate GTP hydrolysis, a key regulatory process.
- This study lays the groundwork for understanding the structural basis of Ras signaling in cellular growth and disease.