Characterizing Viruses with Mass Spectrometry

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The goal of our proposed research is to develop novel approaches for characterizing viruses, initially we will focus on human rhinovirus (HRV), flock house virus (FHV) and adenovirus, and further, use this technology as a general assay for the identification of viruses as well as viral cellular receptors. One approach is the selective extraction of HRV viruses from solution onto a plate using antibodies or nucleic acid reactive tethers followed by analysis of the proteolyzed capsid proteins. Essentially, this would be developed as a two-step assay (binding combined with mass spectrometry) to extract and identify viruses from biofluids. Furthermore, by tethering HRV and adenovirus to a surface we plan to perform affinity experiments with cellular protein extracts with the goal of identifying cellular receptors via protein mass mapping. A more specific outline of our research aims are as follows: 1. Develop affinity approaches attached to a solid support to selectively capture viruses. The target virus systems will be extracted from the solution and identified by proteolytic mass mapping of their capsid proteins. Desorption/ionization techniques such as matrix-assisted laser desorption/ionization (MALDI) and desorption/ionization on silicon (DIOS) will be used initially. Liquid chromatography tandem mass spectrometry will also be used for these analyses to allow for the analyses of heterogeneous protein samples. As there are no rapid, inexpensive screens for most viral infections this approach will be further developed as a general method for identifying viruses. 2. Identify cellular protein receptors using immobilized virus. Whole immobilized adenovirus and HRV will be subjected to cellular extracts, followed by removal of unbound material and subsequent analysis of the proteins bound to the virus. Proteolysis followed by mass analysis and database searching will allow for the characterization of viral receptors.