Select Configurational Dynamics in Ethanolamine Ammonia-Lyase Radical Enzyme Catalysis

Contributions of protein and coupled solvent configurational fluctuations to the molecular mechanism of enzyme catalysis are addressed in the adenosylcobalamin (coenzyme B₁₂)-dependent ethanolamine ammonia-lyase (EAL) enzyme from Salmonella enterica serovar Typhimurium. Full-spectrum, time-resolved electron paramagnetic resonance (EPR) spectroscopy is used to measure the temperature dependence of the first-order kinetics of the substrate radical reaction in EAL surrounded by successive hydration and aqueous-cosolvent (aminoethanol; added dimethyl sulfoxide, glycerol, and sucrose) layers in frozen solution. At different temperature (T) values in each system, the piecewise-continuous Arrhenius relation displays the characteristic bifurcation, from high-T monoexponential dependence (reaction from substrate radical macrostate, S ^• ) to the low-T biexponential dependence (reaction from sequential substates, S ₁ ^• and S ₂ ^• ). Parallel measurements of the solvent dynamics around EAL by using EPR spin probe mobility or electric permittivity detect a dynamical transition from collective cluster to individual fluctuations of coupled protein surface groups and hydration water, with decreasing T, that precisely coincides with the kinetic bifurcation T in each solvent system. When shifted along their common monotonic high-T relation, the Arrhenius dependences collapse onto a single, universal pattern. The results indicate that specific, or select, collective fluctuations in the EAL protein hydration layer are coupled to active-site configurational fluctuations, providing low-barrier portals through the configuration space. The direct mechanistic link between solvent dynamics and turnover expands the understanding of radical-mediated reactions in EAL and supports the model that select collective configurational dynamics are a fundamental feature of enzyme catalysis.