Can enzyme proximity accelerate cascade reactions?

Abstract

The last decade has seen an exponential expansion of interest in conjugating multiple enzymes of cascades in close proximity to each other, with the overarching goal being to accelerate the overall reaction rate. However, some evidence has emerged that there is no effect of proximity channeling on the reaction velocity of the popular GOx-HRP cascade, particularly in the presence of a competing enzyme (catalase). Herein, we rationalize these experimental results quantitatively. We show that, in general, proximity channeling can enhance reaction velocity in the presence of competing enzymes, but in steady state a significant enhancement can only be achieved for diffusion-limited reactions or at high concentrations of competing enzymes. We provide simple equations to estimate the effect of channeling quantitatively and demonstrate that proximity can have a more pronounced effect under crowding conditions in vivo, particularly that crowding can enhance the overall rates of channeled cascade reactions.

Co-localization and confinement of ecto-nucleotidases modulate extracellular adenosine nucleotide pools

“…3) that are accentuated for increasing pore sizes. Hence, as the domain approaches the bulk-like system where AMP can escape from the reaction complex, the advantage of enzyme proximity becomes apparent and is consistent with recent studies of enzyme co-localization [19, 45, 46]. In summary, the nature of the intermediate, AMP, interactions with the surface appear to determine the relative advantage of enzyme colocalization in closed, nanoscale domains.…”

“…More sophisticated models accounting for enzyme size [38–41], charge[42, 43] and co-distribution [44] are based on ordinary and partial differential equation formalisms that implicitly capture these effects. Recent ordinary differential equations (ODE) approaches that implicitly consider the distributions of finite-sized enzymes include a mean field theory from Rao et al and[45]. These models provided strong quantitative insights into the efficiency of catalytic processes [46] and limits on efficiency gains for sequentially-coupled enzymes [45], but only implicitly account for geometrical and physiochemical factors.…”

“…Recent ordinary differential equations (ODE) approaches that implicitly consider the distributions of finite-sized enzymes include a mean field theory from Rao et al and[45]. These models provided strong quantitative insights into the efficiency of catalytic processes [46] and limits on efficiency gains for sequentially-coupled enzymes [45], but only implicitly account for geometrical and physiochemical factors. Explicit consideration of those factors for coupled enzyme processes generally utilize partial differential equations or particle-based solutions, which have afforded descriptions of how neighboring reactive enzymes [29, 47, 48], feedback inhibition, [29, 46], protein geometry and electrostatic interactions [19, 21, 49, 50] contribute to enzyme activity.…”

DNA-Scaffolded Proximity Assembly and Confinement of Multienzyme Reactions

“…The simulation of cascade kinetics showed that proximity did not contribute to the activity enhancement of the assembled GOx-HRP pairs. Other more recent reaction-diffusion modelings suggested that a close proximity between enzymes could only enhance the rate of a cascade reaction under crowding conditions where the diffusion of enzymes and substrates was significantly slowed down [93]. Similar modeling techniques have also been applied to in vivo metabolic pathways, which likewise showed that diffusion was not a rate-limiting factor for many enzyme systems and, therefore, proximity or substrate channeling would not significantly increase the overall rate of the cascade reaction at steady state [88,93].…”

“…Other more recent reaction-diffusion modelings suggested that a close proximity between enzymes could only enhance the rate of a cascade reaction under crowding conditions where the diffusion of enzymes and substrates was significantly slowed down [93]. Similar modeling techniques have also been applied to in vivo metabolic pathways, which likewise showed that diffusion was not a rate-limiting factor for many enzyme systems and, therefore, proximity or substrate channeling would not significantly increase the overall rate of the cascade reaction at steady state [88,93]. Thus, substrate channeling is more likely used to regulate metabolite flux, improve pathway selectivity and protect metabolites from degradation or competing side reactions [88,93].…”