The enzyme Dihydropteridine reductase (DHPR) is found in the brain where it participates in the reproduction of tetrahydrobiopterin from quinonoid-dihydrobiopterin during the hydroxylation cycle of phenylalanine, tyrosine and tryptophan. In this synthesis, tetrahydrobiopterin is a cofactor to the phenylalanine, tyrosine and tryptophan hydroxylases. This hydroxylation process generates the precursors of important neurotransmitters such as L-Dopa, catecholamine and serotonin. A deficiency in DHPR leads to a neurological disease known as phenylketonuria (PKU).By understanding the interactions between DHPR and its ligands, we can manipulate this enzyme in a way that will help us obtain insights into how DHPR works and perhaps device ways of regulating the enzyme. In recent researches, the X-ray crystal structure of the apoenzyme and the cofactor bound enzyme was determined. However, the crystal structure of the enzyme together with its substrate and cofactor are yet to be known.In this experiment, we use fluorescence, Fourier- Transform Infrared spectroscopy, and ab initio calculations to obtain the spectral characteristics of DHPR, NADH and MTX by themselves. In future experiments we will investigate the interaction when either the cofactor or the inhibitor is binded to the enzyme. Finally, we will identify the spectroscopic changes of the enzyme in a ternary complex.