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Analyses of a Select Set of BRAF Mutants and Implications for Their Mechanistic Action as Drivers of Carcinogenesis

  • Dennis M. Bjorklund,   
  • Xavier Jeanne,   
  • Chrisostomos Prodromou *

Received: 09 Aug 2025 | Revised: 27 Sep 2025 | Accepted: 29 Sep 2025 | Published: 11 Oct 2025

Abstract

Oncogenic BRAF mutations represent a variety of amino acid changes at multiple locations within the BRAF kinase domain. Their classification, as class I, II or III, depends on their RAS dependency, their dimerization status in their signalling state and kinase activity. However, many of these mutants have not been fully characterised for their in vitro oligomeric state and kinase activity. Additionally, their interaction with the Hsp90-CDC37 complex, an important component in their stability and activation, is poorly defined. As these properties likely affect the exact mechanism by which these mutants drive carcinogenesis it is of clinical importance that such a characterization is undertaken. Here we report the purification and characterization of a select number of BRAF-mutant proteins so that we can better understand their mode of action. We find that the purified class-II E586K-mutant kinase domain of BRAF is active in its monomeric state, whereas the class-II mutant L597V is, in contrast, an inactive monomer. We also find that purified T599W, where the tryptophan substitution eliminates an activating-phosphorylation site, was active for kinase activity. Consistent with the monomeric state of these mutant BRAF proteins, they interacted with CDC37, although T599W did show somewhat reduced affinity. Our findings, question the strict classification of specific BRAF mutants and suggest further investigations are required to fully understand their carcinogenic mechanism.

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Bjorklund, D. M.; Jeanne, X.; Prodromou, C. Analyses of a Select Set of BRAF Mutants and Implications for Their Mechanistic Action as Drivers of Carcinogenesis. Biomolecular Mechanisms and Innovations 2025, 1 (1), 2.
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