Visualizing Conformational and Compositional Heterogeneity by Single-Particle EM

Friday 6/19
Dipali Shashital
Iowa State University

Macromolecular interactions that dictate biological processes are often highly dynamic and transient, leading to conformational and compositional heterogeneity of functionally relevant complexes. Single-particle electron microscopy (EM) facilitates the structural determination of distinct species present within complex mixtures, enabling the identification of multiple biologically important conformations from a single sample. In collaboration with NRAMM, we have used automated negative-stain EM data collection and single-particle analysis to characterize the structures of particle sub-populations from highly heterogeneous samples. Automation was key to our previous studies, in which we determined the composition and conformation of in vivo ribosome assembly intermediates from Escherichia coli. Similarly, automation has been crucial for our current work, which focuses on determining the structure and oligomeric state of heterogeneous Tec kinase complexes. Tec kinases play essential roles in immune cell activation, and their activity is controlled by intramolecular interactions that may result in the formation of kinase oligomers. Our initial analysis of oligomeric Tec kinase by single-particle EM provides the first structural data for higher-order complexes formed by these enzymes and will be critical in informing our understanding of kinase autoregulation.

 

2018-01-30T21:17:15+00:00June 19th, 2015|Forums|