PIXE-NRA Analysis to Determine Metalloprotein Complete Stoichiometry
Dr. Paul DeYoung and Dr. Graham Peaslee
While approximately a third of all proteins are metalloproteins, their stoichiometric ratios are still largely unknown. Current elemental analysis procedures are capable of determining which metals are in a protein, but it is difficult to determine how many. To address this, an ion beam analysis method is being developed using Particle-Induced X-ray Emission (PIXE) and Nuclear Reaction Analysis (NRA). PIXE determines the elemental composition of the sample and provides a metal-to-metal ratio. However, this is not the desired ratio of metal per protein. NRA uses Rutherford scattering cross-sections to determine an exact ratio of number of atoms to number of proteins. When PIXE and NRA are used in conjunction, it is possible to accurately determine the desired metal-to-protein stoichiometric ratio. In order to ensure accuracy, this method is being developed on Cyanocobalamin (B12) and Cytochrome C which have known stoichiometries. This method will provide us with a standard such that it will be possible to obtain the atomic ratios of additional metalloproteins with confidence. Recent advances include revising sample preparation, refining the substrate, and improving data analysis.
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