How We Probe the Unknown World
At the core, we are structural biochemists. That means we rely on structural information, obtained ourselves or by others, to understand biochemistry (i.e., how biomolecules interact to perform their functions). We rely heavily on NMR spectroscopy as it can provide information on weak interactions, dynamics, and flexibility that other structural techniques do not report on. We also use protein crystallography and, more recently, single particle cryo-EM when the question warrants. Our structural endeavors inform a host of biochemical studies and Klevit group members receive cross-training in multiple areas.
Paradoxically, we have become particularly fascinated by systems that involve intrinsic disorder. A central dogma of structural biology is that protein structure begets function and much of our understanding of the protein structure-function relationship is derived from characterization of stable, well-ordered structures. But up to 40% of the human proteome, including BRCA1 and BARD1, human small heat shock proteins and their clients, exist in flexible, disordered states that have defied efforts to characterize experimentally. We use techniques including NMR and Hydrogen-Deuterium Exchange/Mass Spectroscopy (HDXMS) to obtain fine-grained, residue-level information about intrinsically disordered regions (IDRs) and the “fuzzy” complexes they form. Our first foray into IDRs and fuzzy complexes was a collaborative study on transcriptional activators/coactivators (Brzovic Mol Cell. 2011). We have subsequently identified a Ubiquitin-conjugating (E2) enzyme that functions via an interaction between its own intrinsically disordered region and that of its substrates (Vittal, Nat Chem Biol. 2015). Most recently, we have uncovered what we have called “quasi-order” in the intrinsically-disordered region of a human small heat shock protein, HSPB1 (Clouser eLife 2019).