Dihydroorotate dehydrogenase (DHOD) catalyzes the only redox reaction in the pathway for pyrimidine biosynthesis. In this reaction, a proton is transferred from a carbon atom of the substrate to a serine residue, and a hydride is transferred from another carbon atom of the substrate to a cofactor. The deprotonation of the substrate is postulated to involve a proton relay mechanism along a hydrogen bonding pathway in the active site. In this paper, molecular dynamics simulations are used to identify and characterize potential hydrogen bonding pathways that could facilitate the redox reaction catalyzed by human DHOD. The observed pathways involve hydrogen bonding of the active base serine to a water molecule, which is hydrogen bonded to the substrate carboxylate group or a threonine residue. The threonine residue is positioned to enable proton transfer to another water molecule leading to the bulk solvent. The impact of mutating the active base serine to cysteine is also investigated. This mutation is found to increase the average donor-acceptor distances for proton and hydride transfer and to disrupt the hydrogen bonding pathways observed for the wild-type enzyme. These effects could lead to a significant decrease in enzyme activity, as observed experimentally for the analogous mutant in Escherichia coli DHOD.