Computational studies on tetrahydrobiopterin and tetrahydroneopterin

Includes bibliographical references (p. 63-66).Tetrahydrobiopterin is an important coenzyme involved with all three aromatic amino acid hydroxylases, glycerol ether monooxygenase, and nitric oxide synthase. Its full name is (6R, 1'R, 2'S)-6-(1',2'- dihydroxypropyl)-5, 6, 7, 8- tetrahydrobiopterin. Neopterin is a related pterin, formed as the result of a deficiency in one of the tetrahydrobiopterin synthesis enzymes, PTPS. Neopterin serves as a marker of the cellular immune system activation. If cellular enzymes are capable of reducing neopterin, one product would be tetrahydroneopterin which, because of its similar structure, might interfere with the normal functioning of tetrahydrobiopterin. The structures of both the tetrahydrobiopterin and tetrahydroneopterin are shown below: [molecular diagrams of: Tetrahydrobiopterin (R,R, S) and Tetrahydroneopterin (R,S,R)]. In order to develop new potentially-useful pharmaceuticals and to simply understand the mechanisms of tetrahydrobiopterin (THB) cellular utilization, it is useful to investigate the conformations of these molecules and their interactions with cellular enzymes by computational means. Our research focused on obtaining 3-dimensional conformations of both tetrahydrobiopterin (THB) and tetrahydroneopterin (THN), using the quantum mechanical computational program Gaussian 09, to find the lowest energy state conformations of the molecules studied. We began by performing an overall scan of possible configurations of THB and THN molecules, obtaining the lowest energy conformations for each molecule. Next, we ran the DFT feature of the Gaussian 09 program for further study of the conformations showing the lowest energy conformations. Our work is the first to consider effects of water on THB and THN conformations; we used two approaches. In the first, we added one or two water molecules to THB molecules to observe their effects on hydrogen bonding and structure. In the second, we simulated the effects of solvating our THB and THN molecules using the computer program COSMOtherm. Using the program Discovery Studio 2.5, we tested the ability of THB and THN to fit into the active sites defined by x-ray crystallographic studies of the enzyme structures (docking the THB and THN onto the enzyme). We studied their binding to each of the three aromatic amino acid hydroxylases, The H-bonding patterns were very comparable to the original H-bonding patterns of the THB in the aromatic amino acid hydroxylase structures. Nitric Oxide Synthase (NOS) interacts with THB in a different manner than that with the three aromatic amino acid hydroxylases. In the reaction catalyzed by NOS, THB gets converted into a radical form, either ??BH??? or ??BH??????. We studied both ??BH??? and ??BH?????? using Gaussian 09 and obtained their lowest energy forms. We obtained partial charge and spin density distributions that showed the electron of the ??BH?????? radical was much more delocalized than that of ??BH???, and therefore more stable than ??BH???. Docking of the neutral THB to NOS yielded much better results than docking either ??BH??? or ??BH?????? onto the enzyme active site. We conclude that THB and THN exist in a equilibrium of conformations, varying between the keto and enol tautomers and the axial and equatorial side chain shifts. In both the neutral and the radical form of THB, the keto-axial is the lowest energy form and thus is the predominant conformation. The THB is in its neutral form before binding fully to the active site in the NOS. This is supported from the multiple docking studies showing better overlapping of THB than that of either ??BH??? or ??BH??????. In addition to the above studies, we examined aspects of the recycling pathway, specifically the destruction of THB through the non-enzymatic reaction of the quinoid form of 7,8 dihydrobiopterin (qDHB) formed in the normal recycling reaction to give 7,8 dihydrobiopterin (DHB). We further studied the reaction of DHB to give pterin and 2-hydroxypropanal. Theoretical intermediates were proposed for both reactions. ??Gs were calculated for DHB and intermediates in both the solvated and the unsolvated forms. The results indicated that the two destructive reactions are thermodynamically favore