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

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 tiss...

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Bibliographic Details
Published in:ACS synthetic biology 2020-02, Vol.9 (2), p.191-197
Main Authors: Lee, Jungmin, Vernet, Andyna, Redfield, Katherine, Lu, Shulin, Ghiran, Ionita C, Way, Jeffrey C, Silver, Pamela A
Format: Article
Language:English
Subjects:
Animals
Cell Communication - physiology
Cell Line
Darbepoetin alfa - therapeutic use
Epitopes - genetics
Epitopes - metabolism
Erythrocytes - cytology
Erythrocytes - metabolism
Erythropoietin - genetics
Erythropoietin - metabolism
Glycophorins - immunology
Glycophorins - metabolism
Hemorrhage - drug therapy
Humans
Ligands
Mice
Mice, Transgenic
Mutagenesis, Site-Directed
Protein Engineering
Recombinant Fusion Proteins - biosynthesis
Recombinant Fusion Proteins - isolation & purification
Recombinant Fusion Proteins - therapeutic use
Single-Chain Antibodies - genetics
Single-Chain Antibodies - metabolism
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Summary: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. The 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. On the basis of 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, mesoscale geometry can inform the design of synthetic-biological systems.
ISSN:2161-5063
2161-5063
DOI:10.1021/acssynbio.9b00273