Acquisition of a Structural NMR Resource

点击翻译按钮获取中文摘要

Shared training and research facilities at the Albert Einstein College of Medicine have been developed substantially during the past decade. The focus of these facilities has been in structural and mechanistic biology. These new facilities include a) an x-ray crystallography suite with facilities for three faculty members and trainees, b) a mass spectrometry facility with three instruments, faculty and staff, c) a facility for micro-scale peptide synthesis, isolation, sequencing and analysis, d) a laser spectroscopy laboratory for time-resolved molecular spectroscopy which supports a broad group of faculty and trainees, e) a synchrotron beam-line at Brookhaven National Laboratories which currently supports research and training activities of faculty members in several departments. These new facilities have added to the strengths of existing programs in pulsed EPR spectroscopy (Dr. Jack Peisach), and biological NMR (Dr. Raj Gupta). Together, these facilities make the Albert Einstein College of Medicine one of the premier training sites for structural biology in the greater New York area. Each year, a larger fraction of the approximately 300 Ph.D. and postdoctoral trainees are involved in structural biology training as part of their research programs. A weakness in our training and research programs has been the deficiency in multipleresonance high-field NMR equipment of the type required for structural determination of biological molecules by current technologies. This deficiency was mentioned in the written critiques for each of the past two site-visits for training grants in structural biology and biophysics at this school. This proposal describes a plan to remedy this weakness by the construction of a structural NMR resource containing two spectrometers and associated workstations for molecular modeling: 600 MHz 1H13C15N triple resonance 2.5 and 5 mm probes, 4 channel system, 3-axis gradients 300 MHz 1H13C31P19F 5 mm probe and 10 mm broadband multinuclear probe, 2 channels four 175 MHz UNIX graphics workstations located in the facility for offline data analysis The facility will be newly constructed for the purpose of housing this equipment and will be located on the ground floor of the Ullman Research Building, excavated into foundation bedrock. A highly qualified faculty member, Dr. Mark Girvin, has been recruited to oversee the facility. A full-time staff person will also be provided. The structural NMR resource will operate as a shared research and training facility available to all faculty and their trainees. Examples of research problems under investigation by faculty and their trainees include: structure of the transmembrane subunit c of F1Fo ATP synthase from E. coli melting rates of stem-loop RNA secondary structure and the influence of RNA binding proteins structure/function relationships for cell-surface N-linked complex carbohydrates structure of domains which constitute protein-localization binding sites structure of chromogranin A1-40, a disulfide secretory protein and the effect of AA-replacements structure of the linker region of the membrane protein P-glycoprotein determination of catalytic site flap movements by NMR of 15N-labeled tyrosine phosphatase analysis of the binding of carbohydrates to lectins by NOE and paramagnetic effects In addition to fostering these research programs, the structural NMR resource will be used as the site for a hands-on advanced level graduate school course in the theory and practice of solving structural problems by NMR. The course is to be taught every two years so all Ph.D. students will have the opportunity to learn NMR methods of structural analysis. The location of the graphics workstations in the structural NMR resource will facilitate use of the laboratory for graduate an d postgraduate training.