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Elucidating structure‐function relationships of the human protein kinase MEK1
Author(s) -
Lee Rebecca T.,
Ravatt Leandre M.,
Runco Caroline E.,
Oza Javin P.
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.662.14
Subject(s) - phosphorylation , microbiology and biotechnology , signal transduction , protein kinase a , kinase , biology , mapk/erk pathway , population , demography , sociology
MEK1 is a dual specificity protein kinase in the MAPK phosphorylation cascade. As one of the prevalent signal transduction pathways, the MAPK pathway regulates a diverse variety of cellular processes including proliferation, differentiation, and apoptosis. The phosphorylation event between MEK1 and its only known substrate ERK2 acts as regulatory bottleneck for signal transduction, and maintains the fidelity of signals coming in and going out. As such, MEK1 remains a target for therapeutic intervention of a variety of cancers. While much is known about MEK1's role in cell signaling, enzymatic characterization of MEK1 has been limited due to difficulty in obtaining the active, doubly‐phosphorylated MEK1 in a homogenous population. Our study overcomes this barrier by utilizing an expanded genetic code for the co‐translational incorporation of phosphoserine into MEK1. Specifically, we are leveraging this capacity for on‐demand production of site‐specifically phosphorylated MEK1 to characterize the human MEK1 and elucidate the structure‐function relationships that governs MEK1 activity. Using a hypothesis‐driven approach based on conserved kinase motifs, we selected residues within the activation segment and catalytic loop for site‐specific mutagenesis. Here we report our efforts to express and characterize the phosphorylated MEK1 and its mutants through immunoassays for a comparative analysis of kinase activity. Our work aims to establish kinetic parameters for the human MEK1 for the first time and provide insights into mechanisms of substrate binding and catalysis. Support or Funding Information Our research is funded by the Center for Applications in Biotechnology @ Cal Poly and the Bill and Linda Frost Fund. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .