We are attempting to determine the nature of the signals transmitted to the nucleus that result in cell division. Reversible protein phosphorylation appears to be important in regulating some of these early biochemical events. Therefore, the potentially relevant protein kinases, phosphoprotein phosphatases, and their substrates are purified and characterized. This project seeks to determine the nature of the molecular interactions between Erk (extracellular receptor kinase) and Mek (MAPK/Erk kinase) in this signaling pathway, especially the effect of phosphorylation on kinase activation. We have expressed and purified these proteins as native and GST- and His-tag fusion proteins. In addition we have constructed kinase-inactive and -constitutively active mutants, as well as phosphorylation site mutants. We will use these reagents with a novel flow cytometric technique to measure molecular interactions. GST-Erk will be attached to a glutathione labeled bead and the binding of fluorescent Mek will be measured. This approach will give us information on binding affinity and dynamics not available from conventional methods.
| Frumkin, Jesse P; Patra, Biranchi N; Sevold, Anthony et al. (2016) The interplay between chromosome stability and cell cycle control explored through gene-gene interaction and computational simulation. Nucleic Acids Res 44:8073-85 |
| Johnson, Leah M; Gao, Lu; Shields IV, C Wyatt et al. (2013) Elastomeric microparticles for acoustic mediated bioseparations. J Nanobiotechnology 11:22 |
| Micheva-Viteva, Sofiya N; Shou, Yulin; Nowak-Lovato, Kristy L et al. (2013) c-KIT signaling is targeted by pathogenic Yersinia to suppress the host immune response. BMC Microbiol 13:249 |
| Ai, Ye; Sanders, Claire K; Marrone, Babetta L (2013) Separation of Escherichia coli bacteria from peripheral blood mononuclear cells using standing surface acoustic waves. Anal Chem 85:9126-34 |
| Sanders, Claire K; Mourant, Judith R (2013) Advantages of full spectrum flow cytometry. J Biomed Opt 18:037004 |
| Cushing, Kevin W; Piyasena, Menake E; Carroll, Nick J et al. (2013) Elastomeric negative acoustic contrast particles for affinity capture assays. Anal Chem 85:2208-15 |
| Piyasena, Menake E; Austin Suthanthiraraj, Pearlson P; Applegate Jr, Robert W et al. (2012) Multinode acoustic focusing for parallel flow cytometry. Anal Chem 84:1831-9 |
| Austin Suthanthiraraj, Pearlson P; Piyasena, Menake E; Woods, Travis A et al. (2012) One-dimensional acoustic standing waves in rectangular channels for flow cytometry. Methods 57:259-71 |
| Vuyisich, Momchilo; Sanders, Claire K; Graves, Steven W (2012) Binding and cell intoxication studies of anthrax lethal toxin. Mol Biol Rep 39:5897-903 |
| Chaudhary, Anu; Ganguly, Kumkum; Cabantous, Stephanie et al. (2012) The Brucella TIR-like protein TcpB interacts with the death domain of MyD88. Biochem Biophys Res Commun 417:299-304 |
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