Scientific Report 2007

Chemistry

Organic, Organometallic, and Medicinal Chemistry

M.G. Finn, S. Brown, S.-H. Cho, V. Hong, J. Kuzelka, J. Lau, S. Lee, Y.-H. Lim, S. Presolski, V. Rodionov

In addition to our work on biological polyvalency with engineered virus particles, supported by the Skaggs Institute of Chemical Biology, we focus on the development of catalysts and the synthesis of biologically useful structures. Three of these projects are described in the following sections.

Copper-Catalyzed Azide-Alkyne Cycloaddition

We have continued our development of new catalysts for the azide-alkyne cycloaddition reaction, which has become a principal tool for the synthesis of possible drugs, dendrimers, polymers, and functionalized surfaces in laboratories around the world. Using quantitative kinetics measurements and reaction calorimetry, we screened more than 200 candidate compounds as catalysts this past year. We found that tris(benzimidazole-methyl)amine ligands are strong accelerators of the cycloaddition, allowing the use of small amounts of catalyst for the quantitative conversion of complex azides and alkynes into linked triazoles. A mechanistic investigation revealed that the reaction is quite complex, but several key hypotheses have emerged. Figure 1 shows one of the best new ligands, which bears an alkylcarboxylate group on each of the benzimidazole arms.

Although the ligand is tetradentate, the acetylide complex (compound 1 in Fig. 1) is bound by 3 of the 4 ligand-binding atoms, leaving 1 benzimidazole 'arm' free. A second arm can be released to allow the organic azide to bind in structure 2. Kinetics measurements indicate that the reaction requires 2 copper centers, and so π-coordination to the alkyne by a second metal center is proposed to speed the bond-forming event, leading to intermediate 3. The pendant carboxylate arm may assist in the last step of the process, hydrolysis of the copper-carbon bond to release the product triazole. A complex such as 4 may result, which can be quickly converted back to the starting acetylide 1, completing the catalytic cycle. To further improve the reaction, we are testing this mechanistic scheme by constructing a variety of new ligands and substrates.

Fig. 1. Proposed mechanistic details of the copper-catalyzed azide-alkyne cycloaddition reaction involving a new benzimidazole-based ligand.

Synthesis of New Electron-Dense Labeling Reagents for Structural Biology

Our work with viruses has highlighted a need for new reagents that contain clusters of heavy atoms and reactive functional groups that allow for site-specific attachment to proteins. Such compounds are of great use in labeling complex structures to aid in structure determination by electron cryomicroscopy. In collaboration with the automated molecular imaging group at Scripps Research, we have developed reagents such as compounds 1 and 2 in Figure 2. These compounds contain 3 and 6 osmium atoms, respectively, as well as an N-hydroxysuccinimide ester group for attachment to lysine residues. Derivatives bearing maleimides (for labeling cysteine) and alkynes (for labeling azide-containing unnatural amino acids) are also in hand. Currently, we are evaluating such compounds in electron cryomicroscopy analyses of cowpea mosaic virus, bacteriophage Qβ, and other proteins from collaborators.

Fig. 2. Examples of electron-dense labeling reagents for electron cryomicroscopy.

Inhibitors of Multidrug-Resistance Enzymes

Membrane-bound proteins of the ATP-binding lipid flippase family are often mutated in cancer cells and result in removal of cytotoxic anticancer drugs before the drugs can work, leading to broad multidrug resistance. Using both structure-guided design, in collaboration with G. Chang and coworkers, Department of Molecular Biology, and in situ assembly of fragments in the target enzymes themselves, we are developing new families of molecules that inhibit these enzymes. Steroidal derivatives that strongly inhibit bacterial (MDR3) and human (P-gp) homologs have been synthesized, as well as several that unexpectedly enhance activity. Both types of compounds are important for an improved understanding of the mechanism of drug transport.

Publications

Bourne, C.R., Finn, M.G., Zlotnick A. Global structural changes in hepatitis B virus capsids induced by the assembly effector HAP1. J. Virol. 80:11055, 2006.

Dìaz, D.D., Sen Gupta, S., Kuzelka, J., Cymborowski, M., Sabat, M., Finn, M.G. Bis(formamidine-urea) cmplexes of Ni^II and Cu^II: synthesis, characterization, and reactivity. Eur. J. Inorg. Chem. 4489, 2006, Issue 22.

Finn, M.G. Emerging high-throughput screening methods for asymmetric induction. In: Chiral Analysis. Busch, K.W., Busch, M.A. (Eds.). Elsevier, Amsterdam, the Netherlands, 2006, p. 79.

Hawker, C.J., Fokin, V.V., Finn, M.G., Sharpless, K.B. Bringing efficiency to materials synthesis: the philosophy of click chemistry. Aust. J. Chem. 60:381, 2007.

Johnson, J.A., Finn, M.G., Koberstein, J.T., Turro, N.J. Synthesis of photocleavable linear macromonomers by ATRP and star macromonomers by a tandem ATRP-click reaction: precursors to photodegradable model networks. Macromolecules 40:3589, 2007.

Le Baut, N., Dìaz, D.D., Punna, S., Finn, M.G., Brown, H.R. Study of high glass transition temperature thermosets made from the copper(I)-catalyzed azide-alkyne cycloaddition reaction. Polymer 48:239, 2007.

Li, C., Finn, M.G. Click chemistry in materials synthesis, 2: acid-swellable crosslinked polymers made by copper-catalyzed azide-alkyne cycloaddition. J. Polym. Sci. A Polym. Chem. 44:5513, 2006.

Prasuhn, D.E., Jr., Yeh, R.M., Obenaus, A., Manchester, M., Finn, M.G. Viral MRI contrast agents: coordination of Gd by native virions and attachment of Gd complexes by azide-alkyne cycloaddition. Chem. Commun. (Camb.) 1269, 2007, Issue 12.

Singh, P., Prasuhn, D., Yeh, R.M., Destito, G., Rae, C.S., Osborn, K., Finn, M.G., Manchester, M. Bio-distribution, toxicity, and pathology of cowpea mosaic virus nanoparticles in vivo. J. Control. Release 120:41, 2007.

Yang, H.B., Das, N., Huang, F., Hawkridge, A.M., Dìaz, D.D., Arif, A.M., Finn, M.G., Muddiman, D.C., Stang, P.J. Incorporation of 2,6-di(4,4′-dipyridyl)-9-thiabicyclo[3.3.1]nonane into discrete 2D supramolecules via coordination-driven self-assembly. J. Org. Chem. 71:6644, 2006.