Development of Cell Permeable Dyes for in-vivo Live Cell Imaging
EMSL Project ID
9791
Abstract
. Our long-term goal is to develop the necessary reagents and technology for the rapid identification of a substantial fraction of the multiprotein complexes in an organism, thereby permitting a systems level understanding of signaling complexes that allow microorganisms to adapt to diverse environmental conditions. Critical to this goal is the development of new affinity reagents and protein tags that permit the rapid identification and validation of protein complexes. To accomplish these goals, we propose to develop multipurpose protein tags and associated affinity reagents, which promise both the ability to isolate and characterize protein complexes as well as the examination of protein-protein interactions in living microbes. We propose to extend the design of affinity reagents to include cell permeable and fluorescent photo-crosslinking reagents that permit efficient stabilization and visualization of bacterial protein complexes. Utilization of novel affinity reagents that become fluorescent upon binding to engineered tags will permit quantitation of expressed proteins and purification and stabilization of protein complexes. Ultimately high-throughput data collection of time-dependent changes in protein complex formation in response to environmental conditions will be available, thus permitting a systems level understanding of how microbes adapt to environmental change.
Project Details
Project type
Exploratory Research
Start Date
2004-07-01
End Date
2005-07-18
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
Chen B, H Cao, P Yang, MUljana Mayer, TC Squier, 2007, "Identification of an Orthogonal Peptide Binding Motif for Biarsenical Multiuse Affinity Probes." Bioconjugate Chem. 18, 1259-1265.
Chen B, LM Markillie, Y Xiong, MU Mayer, TC Squier, 2007,"Increased catalytic efficiency following gene fusion of bifunctional methionine sulfoxide reductase enzymes from Shewanella oneidensis." Biochemistry 46, 14153-14161.
Chen B, MUljana Mayer, and TC Squier. 2005. "Structural Uncoupling between Opposing Domains of Oxidized Calmodulin Underlies the Enhanced Binding Affinity and Inhibition of the Plasma Membrane Ca-ATPase." Biochemistry 44(12):4737-4747.
Chen B, MUljana Mayer, LMeng Markillie, DL Stenoien, and TC Squier. 2005. "Dynamic Motion of Helix A in the Amino-terminal Domain of Calmodulin is Stabilized Upon Calcium Activation." Biochemistry 44:905-914.