- Institute of Molecular Biophysics
- Kasha Laboratory of Biophysics
- Florida State University
- Tallahassee, FL 32306-4380
- phone: (850) 644-4104
- fax: (850) 644-7244
- odonnell@chem.fsu.edu
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I am investigating the interaction of the Adeno-associated virus type 2 (AAV-2) with its primary cellular receptor heparan sulfate proteoglycan (HSPG) using single particle analysis. This technique involves acquiring thousands of images of AAV-2 (with and without bound receptor) vitrified in a thin water layered sample and grouping them into classes based on similarity. If the structure of the specimen is known, reference projections may be created for the classification. The grouped images are then averaged to reduce random noise concurrently increasing the SNR. Data acquisition is performed under low dose with a focal pair protocol, whereby the first image is collect close to focus and the second, further. While poor image contrast hampers particle location in close to focus images, such focal ranges are desirable because they contain higher resolution information. Alternatively, particles collected further from focus are easier to discern, however do not contain as detailed information. This reciprocal relationship engenders the merit of the focal pair method; the greater image contrast in the second image is exploited for particle location and alignment in the fist image. This technique is particularly useful towards specimens with symmetry because one image will have multiple different views (depending on the symmetry) of the object. Subnanometer structures are increasingly being reconstructed by this technique and often PDB subunits are fitted to generate pseudoatomic structures to which α-helices and β-sheets are identifiable.