4zff | pdb_00004zff
Dual-acting Fab 5A12 in complex with VEGF
Structural highlights
Disease[VEGFA_HUMAN] Defects in VEGFA are a cause of susceptibility to microvascular complications of diabetes type 1 (MVCD1) [MIM:603933]. These are pathological conditions that develop in numerous tissues and organs as a consequence of diabetes mellitus. They include diabetic retinopathy, diabetic nephropathy leading to end-stage renal disease, and diabetic neuropathy. Diabetic retinopathy remains the major cause of new-onset blindness among diabetic adults. It is characterized by vascular permeability and increased tissue ischemia and angiogenesis. Function[VEGFA_HUMAN] Growth factor active in angiogenesis, vasculogenesis and endothelial cell growth. Induces endothelial cell proliferation, promotes cell migration, inhibits apoptosis and induces permeabilization of blood vessels. Binds to the FLT1/VEGFR1 and KDR/VEGFR2 receptors, heparan sulfate and heparin. NRP1/Neuropilin-1 binds isoforms VEGF-165 and VEGF-145. Isoform VEGF165B binds to KDR but does not activate downstream signaling pathways, does not activate angiogenesis and inhibits tumor growth.[1] [2] [3] Publication Abstract from PubMedThe development of dual targeting antibodies promises therapies with improved efficacy over mono-specific antibodies. Here, we engineered a Two-in-One vascular endothelial growth factor (VEGF)1 /Angiopoietin 2 (Ang2) antibody with Dual action Fab (DAF) as a potential therapeutic for neovascular age-related macular degeneration (NV AMD). Crystal structures of the VEGF/Ang2 DAF in complex with its two antigens showed highly overlapping binding sites. In order to achieve sufficient affinity of the DAF to block both angiogenic factors, we turned to deep mutational scanning in the complementarity determining regions (CDRs). By mutating all three CDRs of each antibody chain simultaneously, we were able not only to identify affinity improving single mutations but also mutation pairs from different CDRs that synergistically improve both binding functions. Further, insights into the cooperativity between mutations allowed us to identify fold-stabilizing mutations in the CDRs. The data obtained from deep mutational scanning reveal that the majority of the 52 CDR residues are utilized differently for the two antigen binding function and permit, for the first time, the engineering of several DAF variants with sub-nanomolar affinity against two structurally unrelated antigens. The improved variants show similar blocking activity of receptor binding as the high affinity mono-specific antibodies against these two proteins, demonstrating the feasibility of generating a dual specific binding surface with comparable properties to individual high affinity mono-specific antibodies. Deep sequencing-guided design of a high affinity dual-specific antibody to target two angiogenic factors in neovascular age-related macular degeneration.,Koenig P, Lee CV, Sanowar S, Wu P, Stinson J, Harris SF, Fuh G J Biol Chem. 2015 Jun 18. pii: jbc.M115.662783. PMID:26088137[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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