An antibody CDR3-erythropoietin fusion protein.
X-ray crystallographic analysis of a bovine antibody (BLV1H12) revealed a unique scaffold in its ultralong heavy chain complementarity determining region 3 (CDR3H) that folds into a solvent exposed, antiparallel β-stranded "stalk" fused with a disulfide cross-linked "knob" domain. This unusual variable region motif provides a novel approach for generating chimeric antibodies with novel activities. Toward this end, human erythropoietin (hEPO) was substituted for the "knob" domain in this antibody to afford an antibody-hEPO (Ab-hEPO) fusion protein that efficiently expresses in mammalian cells. Ab-hEPO proliferated TF-1 cells with a potency comparable to that of hEPO (EC50 ∼ 0.03 nM) and exhibits a significantly extended plasma half-life (>6 days) in mice relative to hEPO (∼4 h). Mice treated with the Ab-hEPO fusion protein show sustained elevated hematocrit for more than two weeks. This work demonstrates the utility of BLV1H12 CDR3 fusions as a novel approach for generating potent polypeptides with enhanced pharmacological properties.
Department of Chemistry, The Scripps Research Institute , 10550 N. Torrey Pines Road, La Jolla, California 92037, United States.
Yong
Y
Zhang
Danling
D
Wang
Gus
G
Welzel
Ying
Y
Wang
Peter G
PG
Schultz
Feng
F
Wang
D061605:Administration, Intravenous; D000818:Animals; D002417:Cattle; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D018360:Crystallography, X-Ray; D004797:Enzyme-Linked Immunosorbent Assay; D004921:Erythropoietin / Q000494:pharmacology*; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000494:pharmacology*; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000494:pharmacology*
C554270:EPO protein, human; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D004921:Erythropoietin