NiFe hydrogenases are a model system to understand H2 formation and cleavage⁠. H2 is an ideal fuel, since its combustion releases no greenhouse gases, and further understanding of NiFe hydrogenases can lead to improved methods to generate H2 biologically. Hydrogenases have a buried binding site, hidden from solvent. While it has advantages, like substrate selectivity and protection of the catalytic site from inhibitors, like CO, it can impose barriers for H2 to achieve the catalytic site, leading to high Michaelis constants (KM).
 
Computer simulations can be used to study pathways for substrate binding to and unbinding from buried catalytic sites. Simulations have the advantage of considering the motions and flexibility of proteins in solution. Moreover, simulations can also be used to compute association (kon) and dissociation rate constants (koff) for substrate-enzyme complexes, which can be compared to experimental data.

One of the challenges to simulate (un)binding events are the timescales involved: while MD simulations usually achieve the microsecond timescale, (un)binding events can take milliseconds or more to happen. Another challenge is that many (un)binding events must be collected to provide a reliable calculation of kon or koff values. Such challenges are overcome by combining MD simulations with enhanced sampling methods, such as tau-Random Acceleration Molecular Dynamics (tauRAMD)⁠⁠, to increase the chances of observing (un)binding events.

The aims of this project are to characterize the dissociation pathways of substrate and inhibitors fom NiFe hydrogenase and test if mutations in hydrogenases modify such pathways.

NiFe hydrogenase.