Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 107, Issue 8, Pages 3447-3452Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0915057107
Keywords
bovine papilloma virus; E5 protein; expression libraries; protein engineering; traptamers
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Funding
- Yale Center for Excellence in Molecular Hematology (NIH) [DK072442]
- National Cancer Institute (NCI) [CA09159, CA37157]
- Fundacion Alfonso Martin Escudero
- Yale Skin Cancer SPORE [CA121974]
- NIGMS [GM073857]
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This work describes a genetic approach to isolate small, artificial transmembrane (TM) proteins with biological activity. The bovine papillomavirus E5 protein is a dimeric, 44-amino acid TM protein that transforms cells by specifically binding and activating the platelet-derived growth factor beta receptor (PDGF beta R). We used the E5 protein as a scaffold to construct a retrovirus library expressing similar to 500; 000 unique 44-amino acid proteins with randomized TM domains. We screened this library to select small, dimeric TM proteins that were structurally unrelated to erythropoietin (EPO), but specifically activated the human EPO receptor (hEPOR). These proteins did not activate the murine EPOR or the PDGF beta R. Genetic studies with one of these activators suggested that it interacted with the TM domain of the hEPOR. Furthermore, this TM activator supported erythroid differentiation of primary human hematopoietic progenitor cells in vitro in the absence of EPO. Thus, we have changed the specificity of a protein so that it no longer recognizes its natural target but, instead, modulates an entirely different protein. This represents a novel strategy to isolate small artificial proteins that affect diverse membrane proteins. We suggest the word traptamer for these transmembrane aptamers.
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