The concept that catalytic enzymes can act as molecular machines transducing chemical activity into motion has conceptual and experimental support, but much of the claimed support comes from experimental conditions where the substrate concentration is higher than biologically relevant and accordingly exceeds kM, the Michaelis-Menten constant. Moreover, many of the enzymes studied experimentally to date are oligomeric. Urease, a hexamer of subunits, has been considered to be the gold standard demonstrating enhanced diffusion. Here we show that urease and certain other oligomeric enzymes of high catalytic activity above kM dissociate into their smaller subunit fragments that diffuse more rapidly, thus providing a simple physical mechanism of enhanced diffusion in this regime of concentrations. Mindful that this conclusion may be controversial, our findings are sup-ported by four independent analytical techniques, static light scattering, dynamic light scattering (DLS), size-exclusion chroma-tography (SEC), and fluorescence correlation spectroscopy (FCS). Data for urease are presented in the main text and the con-clusion is validated for hexokinase and acetylcholinesterase with data presented in supplementary information. For substrate concentration regimes below kM at which these enzymes do not dissociate, our findings from both FCS and DLS validate that enzymatic catalysis does lead to the enhanced diffusion phenomenon. INTRODUCT
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