News and Publications

Quantum Bioinorganic Chemistry and Photochemistry

* Royal Institute of Technology, Stockholm, Sweden

We use modern methods of quantum chemistry (density functional methods) to investigate the electronic structures, energetics, spectroscopy, and reactions of enzymes and electron-transfer proteins. Much of this research involves transition-metal complexes, particularly iron-sulfur (Fig. 1) and iron-oxo centers, at the active sites of metalloproteins and an electrostatic representation of the complete protein and solvent environment.

Recently, we expanded the scope of our work to a number of new areas. We are examining reaction pathways in natural protein biosensors (photoactive yellow protein) and are characterizing the photophysics of dyes in solution and chromophores in proteins (Fig. 2). We explored the catalytic reaction pathway of tyrosine phosphatases. We also examined how transition-metal ions can catalyze formation of organic heterocycles from smaller multiply bonded precursors ("click chemistry").

The protein electrostatics calculations and analysis are done in collaboration with D. Bashford and his group, Department of Molecular Biology. We are also doing research with them on the energetics of catalytic dephosphorylation by protein tyrosine phosphatases, enzymes important in cell signaling and regulation. We are collaborating with K.B. Sharpless, Z. Demko, V. Fokin, R. Hilgraf, and V. Rostovtsev, Department of Chemistry, on the mechanisms of click chemistry and transition-metal catalysis.

In studies with E. Getzoff and M. Thompson, Department of Molecular Biology, we are investigating photoactive yellow protein. This protein contains a chromophore that undergoes a light cycle and so acts as a protein photosensor. In a related context, following up on experimental work done in the laboratories of R. Lerner, D. Millar, and R. Stevens, Department of Molecular Biology, and in collaboration with F. Salsbury and C. Brooks, also of the Department of Molecular Biology, we closely examined the mechanism of the blue fluorescence of stilbene when this molecule is bound to an antibody. Also, with K. Hahn, A. Toutchkine, and D. Gremyachinskiy, Department of Cell Biology, we are investigating improvements in organic fluorescent dyes, which can act as reporters of protein conformational changes or reactions in cells.

Iron-oxo dimer enzymes catalyze chemically difficult reactions, which are of considerable biological importance. We calculated electronic structures and optimized geometries for active-site structures of the iron-oxo enzymes methane monooxygenase and ribonucleotide reductase in both the oxidized and the 2-electron reduced forms. In evaluations of related energetics in the protein environment, we discovered which surface of the active site is more mobile and reactive and which is anchored more stably to the surrounding protein.

Although the native forms of methane monooxygenase and ribonucleotide reductase have similar active-site structures, the reactions of the 2 enzymes are quite different, involving the oxidation of methane to methanol and the generation of a stable tyrosine radical, respectively. The generation of a stable tyrosine radical is the first step in a radical propagation pathway that results in reduction of ribonucleotides to deoxyribonucleotides, the building blocks for DNA synthesis. We are exploring different models for the activated Fe(III)-Fe(IV) active-site complex called intermediate X in ribonucleotide reductase, which oxidizes tyrosine to the radical form.

Because iron-sulfur proteins are often electron-transfer agents, we seek to understand which features of cluster electronic structure and what interactions with the protein environment determine the wide range of redox potentials in these systems. The study of redox potentials of 4Fe4S proteins revealed the importance of both total cluster charge and hydrogen bonding for establishing redox potentials. The results also indicated how hydrogen bonding stabilizes redox potentials (preferentially stabilizing the more negative, reduced cluster state) in proteins and in synthetic analogs, which have weaker hydrogen bonds than the proteins do.

In a collaboration with J. Fee and C.D. Stout, Department of Molecular Biology, we evaluated the energetic cost of distorting 4Fe4S clusters away from the preferred intrinsic structures. We found that the distortion energy is largest for the cysteine (methylene sulfide) groups and is evidently compensated for by internal strain within the protein and by hydrogen bonding to the cluster cysteines and sulfides.

In previous studies, we showed that in a number of important cases, for example, Rieske iron-sulfur proteins and superoxide dismutases, the redox state and protonation state of the active site are coupled. In addition to being systems of great biological interest, these examples of coupled electron-proton transfer provide lessons for other complex systems, including both iron-oxo enzymes and later stages of the nitrogenase catalytic cycle.

We are continuing electronic structure calculations for the iron-molybdenum cofactor of the nitrogenase enzyme. This cofactor is the 7Fe1Mo active site of the iron-molybdenum protein that binds molecular nitrogen and protons and catalyzes the multielectron reduction of molecular nitrogen to 2 ammonia molecules plus molecular hydrogen. A recent very-high-resolution (1.16 Å) x-ray structure of the "resting form" of the enzyme shows a previously hidden central ligand atom in the prismane cluster. Using a combination of density functional calculations of structures, redox potentials, and Mossbauer isomer shifts, we established the probable oxidation state of the active-site cluster. The central ligand is assigned as a central nitride (N^3-) on the basis of our calculations. This information is a required starting point for studying the catalytic cycle of the enzyme.

Publications

Asthagiri, D., Dillet, V., Liu, T., Noodleman, L., Van Etten, R.L., Bashford, D. Density functional study of the mechanism of a tyrosine phosphatase, I: intermediate formation. J. Am. Chem. Soc. 124:10225, 2002.

Cheng, R.-J., Chen, P.-Y., Lovell, T., Liu, T., Noodleman, L., Case, D.A. Symmetry and bonding in metalloporphyrins: a modern implementation for the bonding analyses of five- and six-coordinate high-spin iron(III)-porphyrin complexes through density functional calculations and NMR spectroscopy. J. Am. Chem. Soc. 125:6774, 2003.

Fee, J.A., Castagnetto, J.M., Case, D.A., Noodleman, L., Stout, C.D., Torres, R.A. The circumsphere as a tool to assess distortion in [4Fe-4S] atom clusters. J. Biol. Inorg. Chem. 8:519, 2003.

Han, W.-G., Lovell, T., Liu, T., Noodleman, L. A density functional evaluation of an Fe(III)-Fe(IV) model diiron cluster: comparisons with ribonucleotide reductase intermediate X. Inorg. Chem. 42:2751, 2003.

Himo, F., Demko, Z.P., Noodleman, L., Sharpless, K.B. Mechanisms of tetrazole formation by addition of azide to nitriles. J. Am. Chem. Soc. 124:12210, 2002.

Himo, F., Demko, Z.P., Noodleman, L., Sharpless, K.B. Why is tetrazole formation by addition of azide to organic nitriles catalyzed by zinc(II) salts? J. Am. Chem. Soc. 125:9983, 2003.

Himo, F., Noodleman, L., Blomberg, M.R.A., Siegbahn, P.E.M. Relative acidities of ortho-substituted phenols, as models for modified tyrosines in proteins. J. Phys. Chem. A 106:8757, 2002.

Lovell, T., Himo, F., Han, W.-G., Noodleman, L. Density functional methods applied to metalloenzymes. Coord. Chem. Rev. 238-239:211, 2003.

Lovell, T., Li, J., Case, D.A., Noodleman, L. FeMo cofactor of nitrogenase: energetics and local interactions in the protein environment. J. Biol. Inorg. Chem. 7:735, 2002.

Lovell, T., Li, J., Noodleman, L. Density functional and electrostatics study of oxidized and reduced ribonucleotide reductase: comparisons with methane monooxygenase. J. Biol. Inorg. Chem. 7:799, 2002.

Lovell, T., Liu, T., Case D.A., Noodleman, L. Structural, spectroscopic, and redox consequences of a central ligand in the FeMoco of nitrogenase: a density functional theoretical study. J. Am. Chem. Soc. 125:8377, 2003.

Lovell, T., Torres, R., Han, W.-G., Liu, T., Case, D.A., Noodleman, L. Metal substitution in the active site of nitrogenase MFe₇S₉ (M = Mo⁴⁺, V³⁺, Fe³⁺). Inorg. Chem. 41:5744, 2002.

Noodleman, L., Lovell, T., Han, W.-G., Liu, T., Torres, R.A., Himo, F. Density functional theory. In: Fundamentals. Lever, B.A. (Vol. Ed.). Elsevier, New York, in press. Comprehensive Coordination Chemistry II: From Biology to Nanotechnology. McCleverty, J., Meyer, T.J. (Eds. in Chief), Vol. 1.

Salsbury, F.R., Jr., Han, W.-G., Noodleman, L., Brooks, C.L. III. Temperature-dependent behavior of protein-chromophore interactions: a theoretical study of a blue fluorescent antibody. Chemphyschem 4:848, 2003.

Thompson, M.J., Bashford, D., Noodleman, L., Getzoff, E.D. Photoisomerization and proton transfer in photoactive yellow protein. J. Am. Chem. Soc. 125:8186, 2003.

Torres, R.A., Lovell, T., Noodleman, L., Case, D.A. Density functional and reduction potential calculations of Fe₄S₄ clusters. J. Am. Chem. Soc. 125:1923, 2003.