Characterizing Inhibition of the CyclinA2-CDK2 Complex by Small Molecular Inhibitors using Two-Dimensional NMR Spectroscopy
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Date
2021-05
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The Ohio State University
Abstract
CyclinA2 is best known for the regulation of the eukaryotic cell cycle through binding and activating CDK2. The role of the CyclinA2-CDK2 protein complex as a regulator of the S and G2 phases of the cell cycle is well characterized, but additional functions of these proteins when acting independently is more enigmatic. While both proteins operate separately in DNA damage repair pathways, CyclinA2 has additionally been implicated in neuronal development and maintenance, working apart from CDK2. Development and application of CyclinA2-specific inhibitors will elucidate the protein's specific roles in the neuron. Prior computational modeling done by Dr. Stephanie Kim identified four potential small molecule inhibitors which were intended to act on an allosteric site of CyclinA2, and were confirmed to be inhibitory with an assay testing the activity of the CyclinA2-CDK2 complex. Our aim was to determine where these inhibitors interact with the CyclinA2-CDK2 complex using 2D 1H-15N NMR spectroscopy. Comparison of the NMR spectra of CDK2 in the presence of ATP and three of the inhibitors indicated that CDK2 interacts with these inhibitors independently of CyclinA2. Furthermore, similarities in the titration spectra of the compounds and ATP suggest that these molecules may act competitively at the ATP binding site. Efforts to determine how CyclinA2 or the CyclinA2-CDK2 complex interacted with the compounds were inconclusive.