Dimerization Induced by C-Terminal 14-3-3 Binding Is Sufficient for BRAF Kinase Activation

Biochemistry. 2020 Oct 20;59(41):3982-3992. doi: 10.1021/acs.biochem.0c00517. Epub 2020 Oct 2.

Abstract

The Ras-RAF-MEK-ERK signaling axis, commonly mutated in human cancers, is highly regulated to prevent aberrant signaling in healthy cells. One of the pathway modulators, 14-3-3, a constitutive dimer, induces RAF dimerization and activation by binding to a phosphorylated motif C-terminal to the RAF kinase domain. Recent work has suggested that a C-terminal "DTS" region in BRAF is necessary for this 14-3-3-mediated activation. We show that the catalytic activity and ATP binding affinity of the BRAF:14-3-3 complex is insensitive to the presence or absence of the DTS, while the ATP sites of both BRAF molecules are identical and available for binding. We also present a crystal structure of the apo BRAF:14-3-3 complex showing that the DTS is not required to attain the catalytically active conformation of BRAF. Rather, BRAF dimerization induced by 14-3-3 is the key step in activation, allowing the active BRAF:14-3-3 tetramer to achieve catalytic activity comparable to the constitutively active oncogenic BRAF V600E mutant.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • 14-3-3 Proteins / chemistry*
  • 14-3-3 Proteins / metabolism*
  • Adenosine Triphosphate / metabolism
  • Catalysis
  • Humans
  • Protein Binding
  • Protein Multimerization
  • Proto-Oncogene Proteins B-raf / chemistry*
  • Proto-Oncogene Proteins B-raf / metabolism*
  • Signal Transduction

Substances

  • 14-3-3 Proteins
  • Adenosine Triphosphate
  • Proto-Oncogene Proteins B-raf