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Mapping of Neutralizing Epitopes on Domain I of the Yellow Fever Virus Envelope Glycoprotein with Recombinant Human Antibodies Generated Through Phage Display
Summary: Dr. ter Meulen is studying the human antibody response to yellow fever.
Yellow fever virus (YFV) is a mosquito-transmitted, enveloped positive-stranded RNA virus belonging to the genus Flavivirus, family Flaviviridae. It causes hemorrhagic fever in humans in Africa and South America. Epidemics of yellow fever (YF) have been occurring with increasing frequency in recent years, with a lethality of about 25 percent. Humoral immunity follows natural YFV infection and immunization with the live attenuated YFV vaccine strain 17D. For humans, the neutralizing epitopes on the virus have not been mapped. In the present study, we constructed two phage libraries displaying human antibody scFv fragments using lymphocytes of two donors who recovered from YF during the 2000 YF outbreak in Guinea, West Africa. To isolate scFvs with specific affinity for YFV, we panned the phage libraries with purified YF 17D virions. Six monoclonal phages with high affinity for the YF 17D virus were isolated. The six different scFv displayed by the monoclonal phages were expressed as soluble molecules and precipitated the YFV envelope glycoprotein (E protein) in a radioimmunoprecipitation assay. Three scFv neutralized YFV 17D in a Plaque Reduction Neutralization Test (PRNT). To map the neutralizing epitopes on the YFV-E protein, viral escape mutants of the 17D YFV strain were generated under selective pressure of the respective scFv. Sequencing analysis of these variants identified single amino acid substitutions on domain I of the E protein at positions aa 153 and 154. Because the epitopes were mapped with recombinant antibodies generated from wild-type YFV–infected patients, we believe that we have identified the domains on the 17D vaccine strain of YFV responsible for inducing protective humoral immunity in humans.
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