Crowding can speed up enzyme reactions—until protein droplets become too packed
Our cells are a hotbed of activity, with reactions churning away to keep us alive. But the cytoplasm, where many metabolic reactions take place, is not uniform. Instead, proteins and other molecules cluster together into membraneless compartments, creating distinct, crowded microenvironments for reactions.
In the cytoplasm of cells, proteins and molecules gather to form dense, crowded microenvironments where many metabolic reactions occur. The precise impact of these crowded conditions on enzymes and their activity, however, is still under research. A recent study by researchers from Okinawa Institute of Science and Technology (OIST), Max Planck Institute for Dynamics and Self-Organization (MPI-DS), and RIKEN has shed light on this topic.
The team created model protein-rich droplets with varying concentrations of a protein called BSA, alongside enzymes lactate dehydrogenase (LDH) and alcohol dehydrogenase (ADH) and their substrates. They observed that, up to a certain concentration, increasing BSA levels enhanced enzyme activity. Nevertheless, at extremely high levels of BSA, crowding negatively influenced enzyme activity, leading to a decrease.
To understand why increased crowding initially boosted enzyme activity, the researchers from RIKEN utilized computer simulations. They discovered that in dilute environments, enzymes have an open active site, allowing substrates to bind and react. However, as the environment became crowded with BSA, the active site closed, lowering the energy barrier required for the reaction to occur. Consequently, the reaction proceeded more readily due to the lower energy barrier.
Next, the researchers examined how enzyme activity affected the fluid dynamics of the droplets. They conducted fluorescent tracer experiments and found that active enzymes made both BSA proteins and fluorescent nanoparticles move more freely within the crowded droplets. Conversely, droplets with inactive enzymes were more viscous, impeding molecule movement and mixing. Furthermore, higher concentrations of enzymes or their substrates also increased droplet mobility.
A theoretical model proposed by MPI-DS researchers suggested that enzyme activity creates chemical gradients within the droplets, inducing subtle flows that enable molecules to move more freely and mix. This discovery highlights the potential for enzyme activity to influence physical processes on larger scales.
The findings suggest that interactions between enzymes and their crowded environment can occur in both directions. Future studies aim to explore whether these two-way interactions hold true for multienzyme complexes, where multiple enzymes are in close proximity, and to conduct experiments in vivo within bacterial cells to determine whether the results hold under more complex biological conditions.
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