MRI: Acquisition of a 300 Megahertz Broadband Nuclear Magnetic Resonance (NMR) for Undergraduate Research and Research Training at Loras College

Loras College, Dubuque IA

Investigators

Abstract

A grant has been awarded to Dr. David Speckhard at Loras College to purchase a high field Nuclear Magnetic Resonance (NMR) spectrometer. Dr. Speckhard and Loras students will use the NMR to study the three dimensional structure of molecules that are used to probe the active sites of enzymes. Enzymes are large proteins that catalyze (accelerate) the chemical reactions that occur in living cells. Each enzyme has a small pocket or cleft, called the active site, where catalysis occurs. The shape of this site determines what kind of reactions they enzyme can accelerate. Probe molecules fit in the site like a hand in a glove. The NMR is used to determine the exact shape of those probes that fit best, thereby revealing the shape of the site. In one case the NMR can be used to evaluate where the site touches the probe giving an even better picture of the site. The enzymes chosen for this study are all involved in cellular energy production, and all bind the phosphate containing compound Adenosine Triphosphate (ATP). Each enzyme can also bind other phosphate contatining compounds unique to their site shape. Examples include pyruvate kinase binding CEPA of PEP, and creatine kinase binding phosphocreatine. In the cell, a metal ion (magnesium) helps the phosphate compounds bind in the enzyme site. Substituting cobalt for magnesium creates a probe molecule that keeps its shpae long enough to study. The NMR is used to detemine which atoms of the probe mjolucule are connected to the cobalt. The NMR will also aid in the study of the glycerokinase site. In this case the probe will be carbon 13 enriched ATP. The MNR will report the distances from the carbon 13 atoms and phosphorous 31 atom in the probe to a manganese atom substituted for magnesium in the enzyme site. These distances can be used to create a good map of the glycerokinase site. The interaction of small molecules with large biomolecules is important in many areas of biochemistry. Studying the shape requirements in several kinase enzymes will help determine how enzymes have the ability to be such powerful and selective catalysts. These methods can be applied to drug receptor interactions to show how new medicines interact with biomolecules inside cells and how they may be designed to be more effective and have fewer side effects. These methods are also useful in agricultural chemistry to help produce better fertilizers and pesticides. A significant feature of the proposal is the opportunity provided to undergraduates to use the high field NMR in their research. This opportunity will significantly improve the educational experience for Loras College students since MNR techniques are so important in their future careers in medicine, biotechnology and chemical research.