Enhanced Diffusion and Oligomeric Enzyme Dissociation.

Area of Science:

• Biochemistry
• Enzymology
• Biophysics

Background:

• Oligomeric enzymes have been proposed as molecular machines converting chemical energy into motion.
• Previous studies on enhanced enzyme diffusion often used substrate concentrations exceeding biological relevance (above kM).

Purpose of the Study:

• To investigate the mechanism of enhanced diffusion in oligomeric enzymes, particularly urease.
• To determine if enzyme dissociation into subunits contributes to enhanced diffusion at substrate concentrations above the Michaelis constant (kM).

Main Methods:

• Utilized four independent analytical techniques: static light scattering, dynamic light scattering (DLS), size-exclusion chromatography (SEC), and fluorescence correlation spectroscopy (FCS).
• Examined enzyme behavior across varying substrate concentrations, focusing on conditions above and below the Michaelis constant (kM).

Main Results:

• Demonstrated that urease and other oligomeric enzymes (hexokinase, acetylcholinesterase, aldolase) dissociate into subunits at substrate concentrations above kM.
• Observed significantly enhanced diffusion for these enzymes only when they dissociated into subunits.
• Found up to 10% enhancement in diffusion coefficient for urease and acetylcholinesterase below kM where dissociation did not occur, aligning with theoretical predictions.

Conclusions:

• Enzyme subunit dissociation above kM provides a simple physical mechanism for enhanced diffusion.
• Enhanced diffusion of oligomeric enzymes is primarily driven by subunit dissociation at supra-Michaelis concentrations.
• Findings reconcile experimental observations with theoretical models of enzyme kinetics and diffusion.