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Rapid Conformational Epitope Mapping of Anti-gp120 Antibodies with a Designed Mutant Panel Displayed on Yeast.*

Posted by on in 2013
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Mata-Fink JKriegsman BYu HXZhu HHanson MCIrvine DJWittrup KD. 2013. J Mol Biol. 425:444-56. doi: 10.1016/j.jmb.2012.11.010. Epub 2012 Nov 15.

Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology and Harvard, Boston, MA 02129, USA.

Abstract

gp120 is a substrate for protein engineering both for human immunodeficiency virus (HIV) immunogen design and as a bait for isolating anti-HIV antibodies from patient samples. In this work, we describe the display of a stripped core gp120 on the yeast cell surface. Validation against a panel of neutralizing antibodies confirms that yeast-displayed gp120 presents the CD4 binding site in the correct conformation. We map the epitope of the broadly neutralizing anti-gp120 antibody VRC01 using both a random mutagenesis library and a defined mutant panel and find that the resultant epitope maps are consistent with one another and with the crystallographically identified contact residues. Mapping the VRC01-competitive antibodies b12 and b13 reveals energetic differences in their epitopes that are not obvious from existing crystal structures. These data suggest mutation sets that abrogate binding to broadly neutralizing antibodies with greater specificity than the canonical mutation D368R, useful in rapidly assessing the nature of a vaccine response.

Copyright © 2012 Elsevier Ltd. All rights reserved.

*The Chicken anti-CMYC used in this publication is a product of Gallus Immunotech Inc.

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