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New activated RAS2 mutations identified in Saccharomyces cerevisiae
B A Wilson1, M Khalil, F Tamanoi
1Department of Molecular Microbiology and Immunology, University of Missouri School of Medicine, Columbia 65212.
Abstract:
Activating mutations in RAS proto-oncogenes encode proteins with greater GTP binding. Such mutant proteins are responsible for many human cancers. Six new amino acids were discovered that can yield an activated Saccharomyces cerevisiae RAS2 protein when they are altered. These new RAS2 alleles were found among a collection of 35 random mutations that exhibit a dominant reduction of glycogen accumulation. The RAS2-P41S and RAS2-E99K alleles encode proteins that have lost responsiveness to GTPase activating proteins. They affect amino acids in loop 2 and helix 3 respectively and illustrate that GTPase activating proteins recognize a larger portion of the RAS structure than previously realized. RAS2 mutations E130K, S153F, A154T, and A157S alter amino acids proximal to the guanine binding site and probably influence nucleotide binding either directly or indirectly.
Insights
New research identified six amino acid changes that activate the Saccharomyces cerevisiae RAS2 protein, offering insights into cancer-related RAS mutations and GTPase activating protein interactions.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Activating mutations in RAS proto-oncogenes are implicated in numerous human cancers.
- RAS proteins bind guanosine triphosphate (GTP) and regulate cellular processes.
Purpose of the Study:
- To identify novel mutations that activate the Saccharomyces cerevisiae RAS2 protein.
- To investigate the structural basis of RAS protein activation and regulation by GTPase activating proteins (GAPs).
Main Methods:
- Generated 35 random mutations in the RAS2 gene.
- Screened for mutations resulting in a dominant reduction of glycogen accumulation.
- Sequenced and characterized identified RAS2 alleles.
Main Results:
- Discovered six novel amino acid substitutions that activate RAS2.
- Identified RAS2-P41S and RAS2-E99K alleles, which are unresponsive to GAPs, affecting loop 2 and helix 3.
- Found mutations E130K, S153F, A154T, and A157S near the guanine binding site, potentially affecting nucleotide binding.
Conclusions:
- These findings expand the understanding of RAS protein activation mechanisms.
- The study reveals that GAPs interact with a larger surface area of RAS than previously thought.
- Identified mutations provide new tools for studying RAS signaling pathways and their role in diseases like cancer.