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Title: Molecular dynamics simulations reveal the conformational dynamics of Arabidopsis thaliana BRI1 and BAK1 receptor-like kinases

item MOFFETT, ALEXANDER - University Of Illinois
item BENDER, KYLE - University Of Illinois
item Huber, Steven
item SHUKLA, DIWAKER - University Of Illinois

Submitted to: Journal of Biological Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/30/2017
Publication Date: 5/30/2017
Citation: Moffett, A.S., Bender, K.W., Huber, S.C., Shukla, D. 2017. Molecular dynamics simulations reveal the conformational dynamics of Arabidopsis thaliana BRI1 and BAK1 receptor-like kinases. Journal of Biological Chemistry. doi: 10.1074/jbc.M117.792762.

Interpretive Summary: Receptor kinases are proteins embedded in the plasma membrane with a portion of the protein outside the cell, a transmembrane domain, and a portion inside the cell. These proteins function to transmit chemical signals from the outside to the inside of the cell to regulate growth and development. Binding of the chemical signal molecule to the extracellular domain of the receptor kinase results in activation of the cytoplasmic protein kinase domain, which then initiates a signal transduction cascade. Two of the most studied plant receptor kinases are known as BRI1 and BAK1, which function together in brassinosteroid signaling. Activation of the BRI1 and BAK1 protein kinase domains is known to involve phosphorylation but other mechanisms have not been investigated. Therefore, we conducted molecular dynamic simulations of the phosphorylated, active protein kinase domains of these proteins starting with previously determined static X-ray crystal structures. The simulations revealed an unexpected tendency of a critical structural helix, known to be essential for eukaryotic protein kinase activity, to unfold suggesting that plant receptor kinase activity may be regulated by structural factors in addition to phosphorylation.

Technical Abstract: Initiation of the brassinosteroid (BR) signaling pathway in plants, which is critical for control of growth and development, occurs through the ligand-induced association of BR-insensitive 1 (BRI1) and BRI1-associated kinase 1 (BAK1), receptor-like kinases on the plasma membrane. While a great deal is known about the general mechanism of activation of these two receptors, the atomic details largely remain elusive due to lack of structural information. In order to extend atom-by-atom understanding of how BR signaling is initiated, we performed all-atom molecular dynamics simulations of the fully phosphorylated BRI1 and BAK1 kinase domains, discovering a great deal of disorder in their alpha-C helices, indicating that both are in sub-optimally active states. We created an in silico chimeric BAK1-Src kinase domain by replacing the native BAK1 alpha-C helix with the stable hSrc alpha-C helix and found that our chimera remained in the active state, suppressing alpha-C helix disorder. We performed circular dichroism experiments on the BRI1 alpha-C helix peptide, verifying the presence of disorder, and analyzed the entire known A. thaliana kinome for alpha-C helix disorder using bioinformatic methods, predicting alpha-C helix disorder in multiple kinase families. We believe that disorder in the alpha-C helix of BRI1 and particularly BAK1 could be a regulatory mechanism for the BR signaling pathway, where a disorder to order transition may occur upon association of BRI1 and BAK1, triggering activation in an analogous fashion to the human epidermal growth factor receptor.