XMRV Protease Plasmid
The clones for XMRV protease, as well as clones for all of the other protein products encoded by XMRV, were produced by Protein Expression Laboratory (PEL) to support an NCI effort to develop standard reagents to study this newly discovered human virus implicated in prostate cancer and chronic fatigue syndrome. In the case of XMRV protease, there were multiple clones produced and this was a critical asset to this recent paper. “It was through using an MBP-protease fusion protein that we had the first indication that the lab was working with an enzymatically active protein,” says Dr. Wlodawer. The structural findings suggest that the XMRV protease may be a member of an evolutionary distinct branch of aspartic proteases.
The clone development project in the PEL was headed by Dr. Dom Esposito with technical support from Vanessa Wall and Jennifer Mehalko. “The project to clone all the XMRV-encoded protein products was an NCI priority in the laboratory” says Dr. Esposito. The cloning work took less than 4 weeks to carry out, and generated more than 40 different expression clones for use in producing protein from multiple host organisms ranging from bacteria to mammalian cells. The clones produced by the PEL have been used in a variety of ways in addition the protein structure work of Dr. Wlodawer’s laboratory. Proteins were purified at the PEL for use in development of serological assays for XMRV infection, and were also used to generate monoclonal antibodies. In addition, clones for various proteins were distributed to researchers across the country for biochemical studies, and are now freely available from the AIDS Reference Reagent Repository. These clones were generated using the Gateway recombinational cloning system, which allows very simple transfer of the protein-coding regions to different vectors for many different purposes.