This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.ABSTRACT:Cubic membranes are symmetric, periodic structures that occur in a wide variety of living systems. There are several types of crystal structure proposed for the observed membrane arrays in a variety biological systems. In previous work (Deng, et al. Protozoology), a template matching method was developed to indirectly identify the crystal type using electron microscopy. The system studied was mitochondria of starved amoeba. Subsequently, we used the same material to determine the crystal type from electron tomographic reconstruction, in comparison to a computer-generated model volume of a perfect crystal of the appropriate type. This validated the template-matching method. While Dr. Deng was a postdoc at the RVBC, we made reconstructions of mitochondria representing examples of two different crystal types, the double-diamond and the gyroid. Extensive use of computer visualization and modeling methods enabled us to not only determine the crystal type (double-diamond any gyroid) but also to understand how the cubic membranes are integrated into the architecture of the mitochondrion, giving insights into their function. We found that the spaces in the cubic area were intracristal space, that was connected to the space between the inner and outer mitochondrial membranes by tubular connecting regions as seen in the other types of mitochondria we have studied. The matrix material was narrowly confined between the membranes in the cubic phase, thus reversing the usual situation in which most of the volume inside the mitochondria is matrix material. Two double-tilt reconstructions of canary cardiac muscle mitochondria after 48h starvation were made, as well as one reconstruction from control canary cardiac muscle. In addition, a reconstruction of mitochondria in dog heart was made. Considerable visualization work was done with mitochondria in reconstructions from the canary specimens, with special emphasis on mitochondria with zig-zag cristae. Dr. Zakaria A.M. Almsherqi from Dr. Dengs lab visited the RVBC for several days to learn about and assist with visualization. Good results were obtained.
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