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Open AccessBrief Report

Allosteric Kinase Inhibitors Reshape MEK1 Kinase Activity Conformations in Cells and In Silico

1
Institute of Biochemistry and Center for Molecular Biosciences Innsbruck, University of Innsbruck, Innrain 80/82, 6020 Innsbruck, Austria
2
Bioinformatics Institute (BII), Agency for Science Technology and Research (A*STAR), Singapore 138671, Singapore
3
Tyrolean Cancer Research Institute, Innrain 66, 6020 Innsbruck, Austria
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Academic Editor: Daniel Abankwa
Biomolecules 2021, 11(4), 518; https://doi.org/10.3390/biom11040518
Received: 15 January 2021 / Revised: 26 March 2021 / Accepted: 27 March 2021 / Published: 30 March 2021
Mutations at different stages of the mitogen-activated protein kinase (MAPK) signaling pathway lead to aberrant activation of the involved protein kinase entities. These oncogenic modifications alter signal propagation which converge on the gatekeeper kinases MEK1/2, transmitting the input signal to ERK1/2. Thus, targeted MEK inhibition causes qualitative alterations of carcinogenic MAPK signals. Phosphorylation of the MEK1 activation loop at the positions S218 and S222 by RAF kinases triggers the conformational alignment of MEK’s catalytic pocket to enable ATP-binding and substrate phosphorylation. We have extended a kinase conformation (KinCon) biosensor platform to record MEK1 activity dynamics. In addition to MEK phosphorylation by BRAF, the integration of the phosphorylation-mimetic mutations S218D/S222D triggered opening of the kinase. Structural rearrangement may involve the flexibility of the N terminal MEK1 A-helix. Application of the allosterically acting MEK inhibitors (MEKi) trametinib, cobimentinib, refametinib, and selumetinib converted activated MEK1 KinCon reporters back into a more closed inactive conformation. We confirmed MEK1 KinCon activity dynamics upon drug engagement using the patient-derived melanoma cell line A2058, which harbors the V600E hotspot BRAF mutation. In order to confirm biosensor dynamics, we simulated structure dynamics of MEK1 kinase in the presence and absence of mutations and/or MEKi binding. We observed increased dynamics for the S218D/S222D double mutant particularly in the region of the distal A-helix and alpha-C helix. These data underline that MEK1 KinCon biosensors have the potential to be subjected to MEKi efficacy validations in an intact cell setting. View Full-Text
Keywords: RAF; RAS; ERK; kinase inhibitor; allosteric inhibitor; cancer mutations; MAPK pathway; RAS–RAF–ERK pathway; personalized therapy; structure simulations; structure dynamics; biosensor; protein-fragment complementation assay; KinCon reporter RAF; RAS; ERK; kinase inhibitor; allosteric inhibitor; cancer mutations; MAPK pathway; RAS–RAF–ERK pathway; personalized therapy; structure simulations; structure dynamics; biosensor; protein-fragment complementation assay; KinCon reporter
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MDPI and ACS Style

Fleischmann, J.; Feichtner, A.; DeFalco, L.; Kugler, V.; Schwaighofer, S.; Huber, R.G; Stefan, E. Allosteric Kinase Inhibitors Reshape MEK1 Kinase Activity Conformations in Cells and In Silico. Biomolecules 2021, 11, 518. https://doi.org/10.3390/biom11040518

AMA Style

Fleischmann J, Feichtner A, DeFalco L, Kugler V, Schwaighofer S, Huber RG, Stefan E. Allosteric Kinase Inhibitors Reshape MEK1 Kinase Activity Conformations in Cells and In Silico. Biomolecules. 2021; 11(4):518. https://doi.org/10.3390/biom11040518

Chicago/Turabian Style

Fleischmann, Jakob; Feichtner, Andreas; DeFalco, Louis; Kugler, Valentina; Schwaighofer, Selina; Huber, Roland G; Stefan, Eduard. 2021. "Allosteric Kinase Inhibitors Reshape MEK1 Kinase Activity Conformations in Cells and In Silico" Biomolecules 11, no. 4: 518. https://doi.org/10.3390/biom11040518

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