Gilbert Reibnegger et al.: Electronic structure of pteridines Pteridines Vol. 10, 1999, pp. 91-94 Electronic Structure of Tetrahydropteridine Derivatives Gilbert Reibnegger1*, Jutta Pauschenwein2 and Ernst R. Werner3 llnstitut fLir Medizinische Chemie und Pregl-Laboratorium, Universitat Graz, Harrachgasse 21 /1I, A-SOlO Graz, Austria 2EDV-Zentrum der Universitat Graz, UniversitatsstraBe 15, A-SOlO Graz, Austria 31nstitut fUr Medizinische Chemie und Biochemie, Universitat Innsbruck, Fritz-Pregl StraBe, A-6020 Innsbruck, Austria (Received September 1, 1999) Summary Recentl); the 4-amino analogue of tetrahydrobiopterin was found to be a strong inhibitor of nitric oxide synthase while being bound to the enzyme in a manner similar to the natural cofactor tetrahydrobiopterin. We were interested in the electronic properties of these and similar compounds and studied therefore the following model tetrahydropteridine structures: tetrahydrolumazine, tetrahydropterin, 4-amino-analogue of tetrahydropterin and N5-methyl-tetrahydropterin. Ab initio quantum chemical computations used the Har­ tree-Fode method with basis set 6-31G** after geometry optimization with basis set 3-21G*. Results reveal dramatic differences in distribution of electronic charge and all the molecular properties derived thereof~ between a) the lumazine system, b ) the normal pterin system, and c) the 4-amino analogue. In contrast, differences of electronic properties between tetrahydropterin and its N5-methyl-derivative are negligible. Our results are compatible with recent speculations that the striking differences between the effects of the tetrahydropterin struculrc and its 4-amino analogue on cnznllatic activity may be due to electronic inter­ action between the pyrimidine moiety of the ptcrin ring systcm and the heme group. Key words: Ab initio, QuanUlm chemistl:~ Visualiz,ltion, Pteridines, Nitric oxide synthase Introduction with the structural characteristics required for tight binding, i.e., the 6R-L-erythro-I,2-dihydroxypropyl In an attempt to invcstigate potential rec~ · ding side chain (I ). As was expected, this compound reactions inside the active nitric oxide synthase proved to be an inhibitor of nitric oxide synthase as dimer, the 4-amino analogue of tetrahydrobiopterin well as dihydropteridine reductase. Remarkably, in was designed. The idea was to combine the struc­ nitric oxide synthase, the 4-amino analogue of tet­ tural feature known to calise inhibition of dihydro­ rahydrobiopterin is capable only of inhibiting stim­ pteridine reductase, i.e. , the 4-amino substitution, ulation by tetrahydrobiopterin but not the basal activity of the enzyme caused by endogenously ~Address for Correspondence and tor Requests for bound tetrahydrobiopterin (I). Thus, complete Reprints: Dr. Gilbert Reibnegger Institut fur Mediz­ inhibition only is achieved when tetrahydrobio­ inisehe Chemie und Pregl-Laboratorium, Harrachgasse pterin-free enzyme is used. 2l/ II, A-8010 Graz, Auso·ia. This srudv is dedicated to Pro­ Tetrahydropteridine derivatives other than tetrahy­ tessor Helmut Wtchter on occasion of his 70th birthday drobiopterin are known to stimulate nitric oxide Pteridines I Vol. 10 I No.3 92 Gilbert Rcibnegger e l al.: Electronic structure of pteridines synthases (2) and recently it has been shown that from a three-dimensional array of data. even the N5-methyl-derivative stimulates the enzyme although showing almost no reaction with oxygen (3). Results Here we show results of quantum chemical cal­ culations on the electronic structure as well as Figure 1 shows the chemical structures of the molecular properties derived thereof, of four model compounds investigated. compounds: tetrahydropterin, tetrahydrolumazine, 4-amino-tetrallydropterin and N 5 -methyl-tetrahy­ a) Mulliken population analysis and partial atomic charges dropterin. The idea was to provide knowledge of the electronic similarities or diflerences between Figure 2 was o btained by a Mulliken population these compounds in order to generate a sound basis analysis and shmvs the atomic partial charges of the for further research on their modes of interactions pyrimidine moiety of the four studied compounds_ with other molecules such as, e.g., an enzyme. Clea rl)~ there are dranlatic difle rences in the charge distribution between the lumazine (Fig. 2a), the Methods Construction of molecular structures Computer represemations of the molecular structures were generated using the program H yperchem Release 4 for Windows (HypeR-ube Inc., Waterloo, Omario) , and their geomeo-ies were optimized using molecular mechanic,; employing the built-in MJ\1 + force field. Ab initio computations All ab initio computations were performed using the quantum chemical program package Gaussian 94 (G94; Gaussian Inc., Pittsburgh, PA) at an INDIGO 2 workstation (Silicon Graphics Austria, Vienna, Austria). Using the optimized geometries figure 1. Chemical structures of the compounds studied. generated by H yperchem in their standard orienta­ a) tetrahydrolwnazine, b) tetrahydropterin, c) 4-amino tion provided by program G94, a Hartree Fock 3- tetrahydropterin, d) N 5-methyl tetrahydropterin. 21G* geometry optimization was performed. With the optimized geometries, a 6-31 G ** single point a -Ott b -O~6 computation was done to compute the el ectronic properties. Electronic charge density fllllctions and 0.847 II 0.806 II 0.348 H,--,"/ ~/ electrostatic potentials were obtained in a cube with 0 030 0.325 _0 :;--1/11~2 -0844 '1 11 . the following dimensions: space c(x)rdinates x,y, and 1.0~ 0.782 0.318 H ;>9/~ /~9 O~-""""""N /~ z starting from -0.5700 11.111 and extending to -0.792 NI N -0.623 I -O.Hn -0.748 +0.5700 11.111 with a step size of 0.012 11.111 . Thus, a H H 96x96A'96 grid was produced, and the properties 0.33 5 0.337 were computed at each of these 884736 grid points. C 0.316 0.313 d -0.657 H, / H o Visualization of results '" -0.792 0.826 0.624 1 II 0.324 H--... ./~/ -:?~/ The visualization of the resu.lting data fIles con­ 0 012 -0.740 N II 0.OS2 -0.835 I 11- . taining the electronic charge densities or electrostatic }.316 0.839 1 0.693 0.318 H 0 . 964/,~ /--.Q.68 I H , ~~ /~ . ' N/' N '-- potentials at each grid point, was achieved by the N N -0.791 I -0.756 -0.782 I -0.746 H program AVS Express (Advanced Visual Systems H 0.337 Inc., Manchester, U .K. ) using a desktop personal 0 3 computer (pentium, 166 MHz, 64 MB RAM) under figure 2. Partial charges of the atoms in the pyrimidine mo iety of the pterin ring system of d1C compounds Microsoft Windows 95. This program enables one to studied. a) tetrahydrolumazine, b) tetrahydropterin, c) 4- plot, e.g., isodensity surfaces and isolines computed amino tetrahydropterin, d ) N5-methyl tetrahydropterin. Pteridines / Vol. 10 / No. 3 Gilbert Reibneggcr et al.: Electronic structure of pteridines 93 4-amino tetrahydropterin (Fig. 2c) and both, the the 0.03 a.u. isopotential surfaces) to +0.10 a.u. normal tetrahydropterin (Fig. 2b) and its NS­ with equal spacing of 0.01 a.u. methyl analogue (Fig. 2d), but importantly, there is It is easily seen that while tetrahydropterin (Fig. essentially no ditlerence between the latter two com­ 3b) and N5-methyl-tetrahydropterin (Fig. 3d) show pounds. a strikingly similar spatial arrangement of the molec­ ular electrostatic potential, tetrahydrolumazine (Fig. b) ELectron density and moLecuLar eLectrosTatic potential 3a) as well as 4-amino-tetrahydropterin (Fig. 3c) both show very unique structures. The lumazine Figure 3 shows the main results of the quantum compound and the two normal pterins shows, for chemical computations for each of the four com­ example, a very similar arrangement of the molec­ pounds studied. The molecular structures are visu­ ular electrostatic potential in the region of C4, C4a alized by means of the inner isodcnsity surfaces and N5 (in tetrahydropterin the hydrogen at NS is (0.10 a.u.) on which the local molecular potential is abovc the ring plane, in the other compounds the mapped by means of a greyscale: the brighter the hydrogen or the methyl group is below this plane), isodensity surface at a specific site, the more posi­ bur of course, in the region ofNl and C2 the spatial tively charged is the surface at this site, and the distribution is totally different. The 4-amino com­ darker, thc more negatively charged is the smface. pound, in contrast, is again quite different; in par­ (A negative probe charge would be attracted to the tindar, the situation at N3 is unique. bright sites of the surfaces, and would be repelled It is important to remember that the molecular from the dark sites). electrostatic potential governs the way how mole­ As the molecular electrostatic potential is the main cules mutually interact. Based on the results of Fig. force governing interactions of a molecule with 3, therefore, Fig. 4 reiterates the features of the other molecules, it is necessary to view the charac­ molecular electrostatic potential function, visualized teristics of this important molecular property also in by a vector field depicting the gradient vectors of some distance from the molecules. To this end, iso­ tlle potential (in the molecular plane): the arrows lines are drawn surrounding the molecules, lying show the way a negative probe charge would choose within the molecular plane (thin lines denote posi­ if it was positioned at a specific site (the starting tive molecular electrostatic potential, thick lines rep­ points of the arrows), and then left to move freely resent negative potential). The extra black isosur­ in the force field of the molecule. Clearly, the arrows faces denote centers of negative molecular electro­ point away from the centers of negative charge, and static potential (isovalue -0.03 <l.u. ), and the isolines arc directed upon positively charged sites of the go from -0.02 a.u . (these are the lines adjacent to molecule. These vector fields thus nicely demon­ strate in a very direct wa\, the ditlercnccs betwecn a a c d c figure 3.