Structures of macromolecular machines
Multi-protein complexes are necessary for nearly all cellular processes, and understanding their structure is required for elucidating their function. Current high-resolution strategies in structural biology are effective but lag behind other fields (e.g., genomics and proteomics) due to their reliance on purified samples rather than heterogeneous mixtures. We are working on a method combining single-particle analysis by electron microscopy with protein identification by mass spectrometry to structurally characterize macromolecular complexes called (shotgun EM).
The cryo-electron microscopy structure of a type IV-B CRISPR ribonucleoprotein (RNP) complex (Csf) at 3.9-Å resolution, reveals this complex best resembles the type III-A CRISPR Csm effector complex, consisting of a Cas7-like (Csf2) filament intertwined with a small subunit (Cas11) filament, but the complex lacks subunits for RNA processing and target DNA cleavage. Surprisingly, instead of assembling around a CRISPR-derived RNA (crRNA), the complex assembles upon heterogeneous RNA of a regular length arranged in a pseudo-A-form configuration.
We are developing algorithms that (1) sort projections that belong to the same 3D structure in a large mixture of complexes using a graphical framework, (2) identify interaction interfaces between protein pairs using shape reduction and chemical properties and evolutionary couplings.
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All content © David Taylor, Dustin Taylor, Candice Chen 2021