This proposal requests funding for the purchase of a TVIPS 4K x 4K CCD camera for our FEI Tecnai F20 (TF20) electron microscope, with a 1K x 1K Fastscan CCD accessory. Research by the major users of the TF20, including six NIH-funded projects, provide cutting edge, state-of-the-art advances in the fields of bacterial pathogenesis, apoptosis, cardiovascular disease, viral replication, protein biogenesis, and muscle regulation. Minor users broaden our research further with four NIH-funded studies on the glycolytic pathway, nitrogen metabolism in bacteria, and the mitochondrial respiratory pathway. The purchase of a new CCD camera for the TF20 will allow us to collect data efficiently and accurately for the following major projects: 1) comparative studies on the specialization of pathogenic bacterial adhesion pili for colonization of specific organs in their human hosts, 2) studies on the apoptosome, a macromolecular assembly that is critical to the mitochondrial apoptosis pathway, 3) studies on the role of `bad cholesterol'in cardiovascular disease: the structure and function of LDL low density lipoproteins alone and in complex with its binding receptor, LDLR, 4) studies on the role of protein oligomerization for the replication of poliovirus RNA, including studies on the 3Dpol polymerase in isolation and tethered to membranes via the protein 3AB, 5) comparative studies of ribosome-Sec61 and ribosome-SecY complexes that mediate co-translational translocation of nascent proteins in mammals and bacteria, and 6) studies on the regulation of skeletal muscle contraction by thin filaments. The addition of a large format CCD to our existing TEM will allow us to remain at the forefront of our respective fields. The 1K x 1K CCD that is currently on theTF20 is an excellent tool for astigmatism correction, focus adjustment, and for checking a sample after a low dose image has been recorded on film. The requested large format CCD captures images of an area that is ten times larger than our current CCD. This increase in the amount of sample imaged is essential, as the limits imposed by our current CCD exclude the possibility of collecting the large datasets required for high resolution image reconstructions. However, the existing CCD will be a significant asset for routine work that is done on our Philips CM12 electron microscope Thus, an additional key benefit of this proposal will be the transfer of our existing TVIPS 1K x 1K CCD to our Philips CM12 electron microscope, for which no funding is requested. The CM12 has a large user base that currently includes the five major TF20 users and seven additional projects, six of which are NIH-funded. There is also a steady influx of new users. Having a CCD on this microscope will be extremely beneficial both as a training tool and for data collection on the CM12. Public Health Relevance: This proposal requests funding to upgrade our FEI Tecnai F20 electron microscope with a digital imaging system. Research by users of the electron microscope, including 10 NIH-funded projects, provide cutting edge, state-of-the-art advances in the fields of cardiovascular disease, bacterial pathogenesis, cell death (apoptosis), viral replication, muscle regulation, protein biosynthesis, and energy generation. Addition of this hardware will significantly improve the productivity of all these research projects, advancing research toward the prevention or elimination of human disease.
Showing the most recent 10 out of 12 publications
