Links Of Conformational Sampling To Functional Plasticity And Clinical Phenotypes By Single Molecule Studies

The dynamic exploration of conformational states, also termed conformational sampling, is known to govern all aspects of protein behavior from folding to function. The way however it encodes enzyme function - the capacity to accelerate the chemical step and plasticity of accepting structurally diverse substrates - are terra incognita for conventional bulk characterization but can be directly observed by single molecule studies(1-3).To interrogate the functional and structural dynamics of individual enzymes we spatially confine them on arrays of native membrane like systems (liposomes or nanodiscs)(4). In addition to providing a native like environment - minimizing deleterious interactions with hard matter, these systems may act as 3D scaffold for the assembly of multiple regulatory bioelements. Our parallel single molecule readout allows us to directly observe, and quantify the extend of conformational sampling for multiple enzymes (P450 oxidoreductases and lipases) and consequently its dependence on regulatory inputs and mutations(5-7). Comparing the readouts on wild type enzymes with their pathogenic mutants with varying plasticity provides the intricate correlation between extends of conformational sampling and plasticity. Providing the first clues of conformational sampling acting as cue for functional and consequently clinical phenotype may pave the way for controlling plasticity and the de novo design of proteins with tailor made functionalities.1. N. S. Hatzakis, Biophys. Chem. 186, 46 (2014).2. T. Ha, Nat Meth 11, 1015 (2014).3. M. F. Juette et al., Curr. Opin. Chem. Biol. 20, 103 (Jun, 2014).4. N. S. Hatzakis et al., Nat. Chem. Biol. 5, 835 (Nov, 2009).5. N. S. Hatzakis et al., J. Am. Chem. Soc. 134, 9296 (Jun, 2012).6. K. Bavishi, N. S. Hatzakis, Molecules 19, 19407 (Dec, 2014).7. T. Laursen et al., ACS Chem. Biol. 9, 630 (Mar, 2014).