Published online 15 March 2006
J1 mutagenesis |
Fusoselect: cellcell fusion activity engineered by directed evolution of a retroviral glycoprotein
Division of Medical Biotechnology, Paul-Ehrlich-Institut 63225 Langen, Germany 1Biotechnology and Bioengineering Group, Institute for Chemical and Bio-Engineering, Swiss Federal Institute of Technology CH-8093 Zurich, Switzerland
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Received December 5, 2005. Revised December 29, 2005. Accepted February 24, 2006.
Membrane fusion plays a key role in many biological processes including vesicle trafficking, synaptic transmission, fertilization or cell entry of enveloped viruses. As a common feature the fusion process is mediated by distinct membrane proteins. We describe here Fusoselect, a universal procedure allowing the identification and engineering of molecular determinants for cellcell fusion-activity by directed evolution. The system couples cellcell fusion with the release of retroviral particles, but can principally be applied to membrane proteins of non-viral origin as well. As a model system, we chose a
-retroviral envelope protein, which naturally becomes fusion-active through proteolytic processing by the viral protease. The selection process evolved variants that, in contrast to the parental protein, mediated cellcell fusion in absence of the viral protease. Detailed analysis of the variants revealed molecular determinants for fusion competence in the cytoplasmic tail (CT) of retroviral Env proteins and demonstrated the power of Fusoselect.
Present address: Christoph Merten, ISISBidogie chimique, 670 83 Strasbourg, France.