The protein kinase C family (PKC) of enzymes has been implicated in a wide variety of processes, including mitotic progression, angiogenesis, carcinogenesis, and metastasis. The overall goal of this research program is to develop molecular tools to establish the intracellular spatiotemporal dynamics of PKC activity.
Specific aims i nclude: 1) High Affinity PKC lnhibitors. Dr. Lawrence will employ a high throughput synthesis/assay protocol to identify potent peptide-derived PKC inhibitors and then evaluate the efficacy of these inhibitory agents in previously described cell-based systems. 2) Fluorescent Probes of PKC Activity. Dr. Lawrence has prepared and evaluated the first example of protein kinase peptide substrates that display a phosphorylation-induced change in fluorescence in living cells. The synthesis and evaluation of analogous substrates for the atypical/novel PKC isoforms will be explored. Substrates capable of sampling the spatial dynamics of PKC activity will be assessed as well. 3) Photoactivatable PKC Inhibitor's and Substrates. Dr. Lawrence will prepare protein kinase substrates and inhibitors that are quiescent until activated by light. These species should prove useful in evaluating the temporal dynamics of protein kinase activity in living cells. 4) Spatiotemporal Dynamics of PKC Isoform Activity During Mitosis. The activity of PKC isoforms during the various stages of mitosis has not been methodically evaluated. Dr. Lawrence will employ the tools developed in the first three specific aims to sample both where and when PKC mitotic activity occurs.
|Garrett, Sarah C; Hodgson, Louis; Rybin, Andrew et al. (2008) A biosensor of S100A4 metastasis factor activation: inhibitor screening and cellular activation dynamics. Biochemistry 47:986-96|
|Sharma, Vyas; Wang, Qunzhao; Lawrence, David S (2008) Peptide-based fluorescent sensors of protein kinase activity: design and applications. Biochim Biophys Acta 1784:94-9|
|Lee, Jung Hwan; Kumar, Sanjai; Lawrence, David S (2008) Stepwise combinatorial evolution of Akt bisubstrate inhibitors. Chembiochem 9:507-9|
|Dai, Zhaohua; Dulyaninova, Natalya G; Kumar, Sanjai et al. (2007) Visual snapshots of intracellular kinase activity at the onset of mitosis. Chem Biol 14:1254-60|
|Lee, Seung-Yub; Liang, Fubo; Guo, Xiao-Ling et al. (2005) Design, construction, and intracellular activation of an intramolecularly self-silenced signal transduction inhibitor. Angew Chem Int Ed Engl 44:4242-4|