Expanding the role of P-31 NMR in the study of phosphate-acting enzymes

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Bruker is the world leader in nuclear magnetic resonance (NMR) spectrometers, and is keen to expand its**presence in catalysis and enzyme inhibition research. Many pharmaceutical targets are enzymes that**chemically transform phosphate-containing compounds. Twelve kinase inhibitor drugs are already used in**cancer chemotherapy, and >100 more in are clinical trials. Detailed understanding of enzyme mechanisms**greatly aids new inhibitor design: the flu drugs oseltamivir and zanamivir, and the cancer drug forodesine were**all designed after determining the target enzymes' transition state (TS) structures. The TS is the top of the**energetic barrier that enzymes must surmount for a reaction to occur. Enzymes bind TSs extremely tightly,**making TS mimics potent inhibitors. However, determining TS structures is technically demanding; it requires**measuring the effects of isotopic labels on reaction rates, effects that are frequently < 1%. Recent advances in**NMR spectroscopy have brought the instrumentation for these measurements within reach of most labs, but the**methods needed to exploit these new capabilities remain under-developed. We propose a combined physical**chemistry / applied enzymology study to develop methods to use phosphorus-31 (31P) NMR to measure**isotope effects. This represents an opportune combination of expertise, with Bruker Canada providing the**NMR expertise while the Berti lab providing expertise in TS analysis. Bruker Canada will directly benefit**from the development of new methods that will become part of their applications library. Both private sector**and academic labs will be able to take advantage of the expanded repertoire of NMR techniques to determine**TS structures, expanding the utility of NMR spectroscopy for non-specialist labs. Bruker Canada will also**benefit from direct access to sample preparation, HQP and the NMR spectrometers available at McMaster for**both development and demonstrations for potential customers. The proposed NMR experiments are expected**to attract the interest of other academic laboratories working on inhibitor design and reaction mechanisms. The**proposed work will therefore expand Bruker Canada's customer base in both the private and public sectors.