Single-Molecule Enzymology

Abstract

Viewing a movie of an enzyme molecule made by molecular dynamics simulation, we see incredible details of molecular motions, be they changes of the conformation or actions during a chemical reaction. Molecular dynamics simulations have advanced our understanding of the dynamics of macromolecules in ways that would not be deducible from the static crystal structures [1, 2]. Unfortunately, these “virtual movies” do not run long enough, compared to the time scale of milliseconds to seconds in which most enzymatic reactions take place. In recent years, rapid advances in the patch clamp technique [3], atomic force microscopy [4, 5], optical tweezers [6, 7], and fluorescence microscopy [8–11] have permitted making single-molecule “movies” in situ at the millisecond to second time scale. These techniques do not have time resolutions as high as that of molecular dynamics simulations, but their single-molecule sensitivities allow probing of conformational motions, which are otherwise masked in ensemble-averaged experiments. Moreover, chemical reactions can now be observed on a single-molecule basis. For example, enzymatic turnovers of a few motor proteins [12–17] and a few enzymes [18–20] have been monitored in real time.