Rational Design of a Bifunctional AND-Gate Ligand to Modulate Cell-Cell Interactions

Abstract

Protein \"AND-gate\" systems, in which a ligand acts only on cells with two different receptors, direct signaling activity to a particular cell type and avoid action on other cells. In a bifunctional AND-Gate protein, the molecular geometry of the protein domains is crucial. Here we constructed a tissue-targeted erythropoietin (EPO) that stimulates red blood cell (RBC) production without triggering thrombosis. EPO was directed to RBC precursors and mature RBCs by fusion to an anti-glycophorin A antibody V region. Many such constructs activated EPO receptors in vitro and stimulated RBC and not platelet production in mice but nonetheless enhanced thrombosis in mice and caused adhesion between RBCs and EPO receptor-bearing cells. Based on a protein-structural model of the RBC surface, we rationally designed an anti-glycophorin/EPO fusion that does not induce cell adhesion in vitro or enhance thrombosis in vivo. Thus, meso-scale geometry can inform design of synthetic-biological systems.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=138 SRC=\"FIGDIR/small/711549v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (22K):\norg.highwire.dtl.DTLVardef@93b382org.highwire.dtl.DTLVardef@ee37ccorg.highwire.dtl.DTLVardef@1913fe4org.highwire.dtl.DTLVardef@f998b1_HPS_FORMAT_FIGEXP M_FIG C_FIG