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Related Concept Videos

Small GTPases - Ras and Rho01:24

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GTPases and their Regulation02:14

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Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒  small G-proteins consisting of a single domain and large multi-domain G-proteins.
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Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high...
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The Ras Gene02:38

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The Ras-gene-encoded proteins are regulators of signaling pathways controlling cell proliferation, differentiation, or cell survival. The Ras-gene family in humans constitutes three primary members—the HRas, NRas, and KRas. These genes code for four functionally distinct yet closely related proteins—the HRas, NRas, KRas4A, and KRas4B. The involvement of mutant Ras genes in human cancer was first discovered in 1982 and is among the most common causes of human tumorigenesis.
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Rab Proteins01:14

Rab Proteins

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Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
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When a ligand binds to a cell-surface receptor, the receptor's intracellular domain changes shape, which may either activate its enzyme function or allow its binding to other molecules. The initial signal is amplified by most signal transduction pathways. This means that a single ligand molecule can activate multiple molecules of a downstream target. Proteins that relay a signal are most commonly phosphorylated at one or more sites, activating or inactivating the protein. Kinases catalyze...
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Comparing the Affinity of GTPase-binding Proteins using Competition Assays
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Comparing the Affinity of GTPase-binding Proteins using Competition Assays

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Allosteric site variants affect GTP hydrolysis on Ras.

Christian W Johnson1, Susan K Fetics2, Kathleen P Davis2

  • 1Department of Chemistry and Chemical Biology, Northeastern University, Boston, Massachusetts, USA.

Protein Science : a Publication of the Protein Society
|August 24, 2023
PubMed
Summary
This summary is machine-generated.

Manipulating the allosteric site of HRAS GTPase impacts GTP hydrolysis. Allosteric site closure increases HRAS GTP hydrolysis, while RAF kinase binding inhibits it, potentially stabilizing the GTP-bound state in cancer.

Keywords:
HRASKRASRAFallosteric siteallosteryhydrolysismutationsmall GTPase

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Cell Signaling

Background:

  • RAS GTPases are key regulators of cell growth and proliferation, cycling between GDP- and GTP-bound states.
  • GTP hydrolysis by HRAS is crucial for signal termination and is potentially regulated by an allosteric site.

Purpose of the Study:

  • To investigate the functional connection between HRAS intrinsic GTP hydrolysis and the allosteric site.
  • To determine how manipulating the allosteric site affects HRAS GTPase activity.

Main Methods:

  • Site-directed mutagenesis to introduce hydrophobic mutations in the HRAS allosteric site.
  • X-ray crystallography to determine structural changes in HRAS mutants.
  • Enzymatic assays to measure GTP hydrolysis rates in solution.

Main Results:

  • Closure of the allosteric site in HRAS mutants correlated with increased intrinsic GTP hydrolysis.
  • Binding to RAF-RBD inhibited GTP hydrolysis in these HRAS mutants.
  • Mutant behavior may mimic tumor-associated mutations stabilizing the GTP-bound RAS state.

Conclusions:

  • The allosteric site and intrinsic GTP hydrolysis of HRAS are functionally linked.
  • RAF kinase binding can counteract the effects of allosteric site mutations on GTP hydrolysis.
  • Understanding these mechanisms may offer insights into RAS-driven oncogenesis.