Research Program

The major thrust of our research program is at the interface of organic, inorganic, and biological chemistry. Many biochemical transformations, as well as important synthetic and industrial processes, are catalyzed by metals.  Here we describe some representative projects of the wide scope of our research in the bioinorganic and bioinspired catalysis fields.  

Porphyrin Catalysts for Organic Transformations

Biochemistry manifests numerous highly selective transformations of aliphatic C-H bonds into alcohols, halides, and olefins catalyzed by reactive metal-oxo intermediates within enzymes. Using this inspiration, we have worked to develop porphyrin catalytic systems to perform these desired synthetic transformations.

Liu, W.; Huang, X.; Cheng, M.-J.; Nielsen, R. J.; Goddard, W. A; Groves, J. T. Oxidative aliphatic C-H fluorination with fluoride ion catalyzed by a manganese porphyrin. Science, 2012, 337, 1322–5.

Liu, W.; Huang, X.; Cheng, M.-J.; Nielsen, R. J.; Goddard, W. A; Groves, J. T. Oxidative aliphatic C-H fluorination with fluoride ion catalyzed by a manganese porphyrin. Science, 2012, 337, 1322–5.

A highly sought after transformation is aliphatic C-H bond fluorination because there is an enormous impetus today to place F at such inaccessible sites in biomolecules and drug candidates. However, biochemistry provides no way to selectively and directly incorporate fluoride ions into unreactive sp3 C-H bonds   A strategy was suggested to us by our recent discovery of a C-H chlorination protocol using manganese porphyrin catalysts and hypochlorite ion. We have discovered that a manganese porphyrin complex catalyzes alkyl fluorination by fluoride ion under mild conditions in conjunction with stoichiometric oxidation by iodosylbenzene. Simple alkanes, terpenoids, and even steroids were selectively fluorinated at otherwise inaccessible sites in 50 to 60% yield. Mechanistic analysis suggests that the regioselectivity for C-H bond cleavage is directed by an oxomanganese(V) catalytic intermediate followed by F delivery via an unusual manganese(IV) fluoride that has been isolated and structurally characterized. Given that the source of F in this one-step, one-pot protocol is fluoride ion, we anticipate the potential application of these techniques to the incorporation of 18F into a wide variety of biomolecules and synthetic building blocks.

Biomimetic Catalysts and Model Systems

The heme prosthetic group is found in a variety of enzymes involved in oxygen metabolism such as the cytochromes P450 (CYP450) of lung, liver and epithelial tissue.  The characterization of synthetic oxo-metalloporphyrin complexes as models of the heme active site has begun to provide a rational basis for the development of more reactive catalysts. New catalysts have also been developed that expand on the reactivity of the biochemical inspiration, such as the production of chlorine dioxide observed when using a manganese catalytic system as a biomimetic catalyst inspired by the oxygen-producing heme enzyme chlorite dismutase.

Bell, S.; Groves, J.T. A Highly Reactive P450 Model Compound J. Am. Chem. Soc. 2009,131(28), 9640-9641.

Bell, S.; Groves, J.T. A Highly Reactive P450 Model Compound J. Am. Chem. Soc. 2009,131(28), 9640-9641.

Significant advances in heme protein enzymology, the characterization of synthetic model compounds, and computational approaches have pointed to a reactive oxoiron(IV)-porphyrin π-radical cation in the consensus mechanism of CYP450. Compound I model compounds, such as the thoroughly characterized [OFeIV-TMP]+, have exhibited a full range of oxygen transfer reactions. However, the low kinetic reactivity of [OFeIV-TMP]+ has left unanswered just how the protein manages to generate a sufficiently reactive intermediate and what that reactive species is. We reported the detection and kinetic characterization of [OFeIV-4-TMPyP]+, which shows extraordinary rates for C-H hydroxylations and is orders of magnitude more reactive than the well-studied tetramesityl analogues. The interesting question is why synthetic OFeIV-porphyrin π-radical cations should have such a great variability in their intrinsic reactivities. We suggest that the high kinetic reactivity observed here experimentally may result from both a low-lying a2u porphyrin HOMO and facilitated spin-state crossing phenomena in the course of the reaction. More generally, the results suggest that even subtle charge modulation at the heme active site of CYP450 to accomplish a similar tuning of state-crossing energies or lower stability of the porphyrin π-radical cation could result in high reactivity of a cytochrome P450 compound I.

Umile, T.P., Groves, J.T. Dissection of the Mechanism of Manganese Porphyrin-Catalyzed Chlorine Dioxide Generation Inorg. Chem. 2011, 50(20), 10353-10362.

Umile, T.P., Groves, J.T. Dissection of the Mechanism of Manganese Porphyrin-Catalyzed Chlorine Dioxide Generation Inorg. Chem. 2011, 50(20), 10353-10362.

Chlorine dioxide has emerged as the preferred agent for microbial decontamination, for wood pulp bleaching and for the detoxification of sites contaminated by biological warfare agents such as anthrax. Recently, we reported the discovery that ClO2 can be produced rapidly and efficiently using a highly electron-deficient, water-soluble manganese porphyrin catalyst. Key advantages of this system are the mildly acidic to neutral reaction conditions, full conversion of the chlorite starting material, high activity on a solid support, and the fact that ClO2 production occurred within seconds. We have investigated the mechanism of the manganese porphyrin-catalyzed chlorine dioxide production from chlorite ion. This process involves rate-limiting oxidation of the manganese (III) catalyst by ClO2- to afford high-valent manganese species. We argue on kinetic, electrochemical, and thermodynamic grounds that this initial intermediate is the same transdioxoMnVTDMImP species that we have previously observed in reactions of this manganese porphyrin with hypobromite and hydrogen peroxide. The mechanism of this catalytic process now suggests methods for greatly enhancing the rate of ClO2 generation under mild, neutral conditions. Efforts to address the practical implementation of manganese porphyrin-generated ClO2 are under way.

Mechanisms of Metalloenzymes

Our interest in oxidizing enzymes has led us to develop techniques for characterizing and using the oxygenase enzymes found within whole cells. This approach has also enabled us to look directly at the mechanism of action of alkane hydroxylases in new and uncharacterized organisms.

Wang, X.; Peter, S.; Kinne, M.; Hofrichter, M.; Groves, J. T.  Detection and Kinetic Characterization of a Highly Reactive Heme-Thiolate Peroxygenase Compound I J. Am. Chem. Soc. 2012, 10–13.

Wang, X.; Peter, S.; Kinne, M.; Hofrichter, M.; Groves, J. T. Detection and Kinetic Characterization of a Highly Reactive Heme-Thiolate Peroxygenase Compound I J. Am. Chem. Soc. 2012, 10–13.

The fungal peroxygenase AaeAPO (EC 1.11.2.1) from Agrocybe aegerita is a new and highly active heme-thiolate protein. It catalyzes a wide range of H2O2-dependent oxidations including alkane and arene hydroxylations, epoxidations, halogenations, and ether cleavage reactions. AaeAPO functions as a monooxygenase, similar to cytochrome P450, and has been shown to produce human metabolites efficiently from drugs. Our study of AaeAPO supports the formation of a highly reactive AaeAPO oxoiron(IV) porphyrin radical cation intermediate. Reaction kinetics for AaeAPO-I with a variety of substrates have revealed an informative correlation between the C−H BDE and the observed bimolecular rate constants. The large estimated FeIVO−H BDE of 103 kcal/mol for AaeAPO-II is significant with regard to its relationship to the pKa of AaeAPO-II and the one-electron reduction potential of AaeAPO-I. These aspects are under investigation.

Cooper, H. L. R.; Mishra, G.; Huang, X.; Pender-Cudlip, M.; Austin, R. N.; Shanklin, J.; Groves, J. T. Parallel and competitive pathways for substrate desaturation, hydroxylation, and radical rearrangement by the non-heme diiron hydroxylase AlkB J. Am. Chem. Soc. 2012, 134, 20365–75.

Cooper, H. L. R.; Mishra, G.; Huang, X.; Pender-Cudlip, M.; Austin, R. N.; Shanklin, J.; Groves, J. T. Parallel and competitive pathways for substrate desaturation, hydroxylation, and radical rearrangement by the non-heme diiron hydroxylase AlkB J. Am. Chem. Soc. 2012, 134, 20365–75.

The alkane monooxygenase AlkB from Pseudomonas putida is an integral membrane, non-heme iron hydroxylase that initiates the terminal hydroxylation of linear alkanes. We have previously shown that AlkB hydroxylates saturated mechanistic probes such as norcarane and bicyclo[3.1.0]hexane, both in whole cell cultures and in cell-free extracts, via a relatively long-lived (1−170 ns) substrate radical intermediate. By comparing the product distribution of the deuterated norcarane to that of the non-deuterated norcarane, we were able to determine that the desaturation products from norcarane derive from hydrogen abstraction at C3, while reaction at C2 results in hydroxylation and radical rearrangement of the substrate.

Reactions of Metalloproteins

Due to their wide influence throughout biology, our interest in metalloproteins extends beyond the direct action of the metal.  Of particular interest to us is the role of heme protein cytochrome c’s release from the mitochondria in cell apoptosis.  Better understanding cell death could have effects in the research of cancer and other diseases.  Additionally, the pathological modification of metalloproteins, such as the tyrosine nitration of manganese superoxide dismutase, poses an interesting mechanistic problem with great significance.

Beales, P.A.; Bergstrom, C.L.; Groves, J.T.; Vanderlick, T.K. Single Vesicle Observations of the Cardiolipin-Cytochrome c Interaction: Induction of Membrane Morphology Changes Langmuir 2011, 27, 6107-6115

Beales, P.A.; Bergstrom, C.L.; Groves, J.T.; Vanderlick, T.K. Single Vesicle Observations of the Cardiolipin-Cytochrome c Interaction: Induction of Membrane Morphology Changes Langmuir 2011, 27, 6107-6115

Simple biomolecular systems consisting of only a few components are useful tools to elucidate the fundamental interactions and behaviors of the constituent species in a way that is inaccessible when studying complex whole cells or living organisms. We developed heterogeneous giant unilamellar lipid vesicles with spatially localized domains enriched in the lipid of interest such that the differential response of disparate membrane compositions can be observed under identical conditions in response to the additional of external stimuli. Cytochrome c (cyt c) is observed to induce budding and collapse of the cardiolipin(CL)-rich domains. We conclude that this phase extrusion is a nonequilibrium phenomenon driven by nonspecific electrostatic interactions between the anionic membrane domains and the polycationic globular proteins that provokes membrane buckling and subsequent aggregation between different segments of the same membrane. These direct single vesicle observations of the CL-cyt c interaction offer compelling insight into their role and function in the mitochondria of the cell, in particular, stabilization of mitochondrial cristae and assisting the escape of cyt c into the cytosol during apoptosis.

Surmeli, N.B..; Groves, Peroxynitrite Medidates Active Site Tyrosine Nitration in Manganese Superoxide Dismutase. Evidence of a Role for the Carbonate Radical Anion J. Am. Chem. Soc.2010, 132, 17174-17185.

Surmeli, N.B..; Groves, Peroxynitrite Medidates Active Site Tyrosine Nitration in Manganese Superoxide Dismutase. Evidence of a Role for the Carbonate Radical Anion J. Am. Chem. Soc. 2010, 132, 17174-17185.

Protein tyrosine nitration has been observed in connection with numerous human diseases including neurodegenerative conditions, cardiovascular disorders, diabetes, and Alzheimer’s disease. Manganese-superoxide dismutase (MnSOD) is one of the critical enzymes that is nitrated and inactivated in vivo under pathological conditions. MnSOD nitration in vivo could be due to direct reaction with peroxynitrite or via freely diffusing •NO2. Knowing which of these species is the causative agent for protein nitration could have important ramifications in our understanding to the origins of cell damage and the underlying disease. We found clear evidence that the positional selectivities of tyrosine nitration for the peroxynitrite and •NO2 pathways are significantly different. Direct reaction with peroxynitrite has been found to result in the nitration of Tyr34, close to the active site manganese, both with and without added CO2, while reaction with freely diffusing •NO2 results in nitration of the most solvent-accessible tyrosine residues (Tyr9 and Tyr11). We also investigated the changes in activity of MnSOD under different conditions and observed that loss of activity in nitrated MnSOD is exclusively due to nitration of Tyr34. The lack of activity of nitrated MnSOD under disease conditions suggests that nitration of MnSOD observed under oxidative stress is due to peroxynitrite directly or via its reaction with CO2.