Modeling Dynamics in the D‐Amino Acid Oxidase Protein

In human brains the enzyme D‐amino acid oxidase (hDAAO) plays a key role in modulating the degradation pathway of the important signaling molecule D‐serine. The hDAAO enzyme is active as a homodimer and is stereospecific for the oxidative deamination of D‐amino acids to give ammonia and an α‐keto acid, coupled to the reduction of the cofactor FAD. The reoxidation of FAD results in hydrogen peroxide. The deregulation of D‐serine signaling has been linked to schizophrenia susceptibility and certain mutations in the hDAAO enzyme itself are associated with familial amyotrophic lateral sclerosis (ALS). Mutations in the hDAAO enzyme are known to impact enzyme function and stability. We were interested in comparing apo‐enzyme simulations to FAD bound simulations of the wild‐type hDAAO enzyme, a wild‐type pig kidney D‐amino acid oxidase enzyme (pkDAAO), a mutant R279A hDAAO enzyme, and a mutant D31H hDAAO enzyme. Even though there is 85% sequence identity between the two species, hDAAO binds FAD weaker and shows a slower rate of flavin reduction compared to pkDAAO. The R279A mutation exchanges an Arginine for an Alanine at position 279 on both chains of the homodimer. This mutation is close to the FAD binding site and has shown to increase the enzyme's binding affinity for FAD. The D31H mutation exchanges an Aspartate for a Histidine at position 31 on both chains of the homodimer. Even though residue 31 is located far from the FAD binding site, this mutation leads to a higher binding affinity for FAD compared to the human wild‐type. We use molecular dynamic simulations to study how the protein structure changes over time and how the protein dynamics are altered by mutations in the amino acid sequence. Investigating the changes we see at the atomic level in protein structure can help to determine how each mutation can have different effects on enzyme stability or activity.