Single-enzyme approach predicts natural emergence of inhibitor-activator duality

The classical theory of enzymatic inhibition aims to quantitatively describe the effect of certain molecules—called inhibitors—on the progression of enzymatic reactions, but growing signs indicate that it must be revised to keep pace with the single-molecule revolution that is sweeping through the sciences. Here, we take the single enzyme perspective and rebuild the theory of enzymatic inhibition from the bottom up. We find that accounting for multiconformational enzyme structure and intrinsic randomness cannot undermine the validity of classical results in the case of competitive inhibition; but that it should strongly change our view on the uncompetitive and mixed modes of inhibition. There, stochastic fluctuations on the single-enzyme level could give rise to inhibitor-activator duality—a phenomenon in which, under some conditions, the introduction of a molecule whose binding shuts down enzymatic catalysis will counter intuitively work to facilitate product formation. We state—in terms of experimentally measurable quantities—a mathematical condition for the emergence of inhibitor-activator duality, and propose that it could explain why certain molecules that act as inhibitors when substrate concentrations are high elicit a nonmonotonic dose response when substrate concentrations are low. The fundamental and practical implications of our findings are thoroughly discussed. Enzymes spin the wheel of life by catalyzing a myriad of chemical reactions central to the growth, development, and metabolism of all living organisms. Without enzymes, essential processes would progress so slowly that life would virtually grind to a halt; and some enzymatic reactions are so critical that inhibiting them may result in death. Enzymatic inhibitors could thus be potent poisons but could also be used as antibiotics and drugs to treat other forms of disease. Inhibitors have additional commercial uses, but the fundamental principles which govern their interaction with enzymes are not always understood in full, and have yet ceased to fascinate those interested in the basic aspects of enzyme science. The canonical description of enzymatic inhibition received much exposure, but even at the level of bulk reactions its many limitations have already been pointed out. Moreover, and despite rapid advancements in the study of uninhibited enzymatic reactions on the singlemolecule level, the study of inhibited reactions has barely made progress in this direction and is still based, by and large, on what is known in bulk.