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Production of multivalent protein binders using a self‐trimerizing collagen‐like peptide scaffold
Author(s) -
Fan ChiaYu,
Huang ChuanChuan,
Chiu WeiChun,
Lai ChunChieh,
Liou GunnGuang,
Li HsiuChuan,
Chou MinYuan
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.08-111484
Subject(s) - trimer , random hexamer , chemistry , peptide , biophysics , scaffold , scaffold protein , biochemistry , cysteine , linker , microbiology and biotechnology , biology , dimer , signal transduction , medicine , organic chemistry , computer science , biomedical engineering , enzyme , operating system
A class of multivalent protein binders was designed to overcome the limitations of low‐affinity therapeutic antibodies. These binders, termed “collabodies,” use a triplex‐forming collagen‐like peptide to drive the trimerization of a heterologous target‐binding domain. Different forms of collabody, consisting of the human single‐chain variable fragment (scFv) fused to either the N or C terminus of the collagen‐like peptide scaffold (Gly‐Pro‐Pro) 10 , were stably expressed as soluble secretory proteins in mammalian cells. The collabody consisting of scFv fused to the N terminus of collagen scaffold is present as a homotrimer, whereas it exhibited a mixture of trimer and interchain disulfidebonded hexamer when cysteine residues were introduced and flanked the scaffold. The collagenous motif in collabody is prolyl‐hydroxylated, with remarkable thermal and serum stabilities. The collabody erb_scFvCol bound to the extracellular domain of epidermal growth factor receptor with a binding strength ~20‐and 1000‐fold stronger than the bivalent and monovalent counterparts, respectively. The trimeric collagen scaffold does not compromise the functionality of the binding moieties of parental immunoglobulin G (IgG); therefore, it could be applied to fuse other protein molecules to acquire significantly improved targetingbinding strengths.— Fan, C.‐Y., Huang, C.‐C., Chiu, W.‐C., Lai, C.‐C., Liou, G.‐G., Li, H.‐C., and Chou, M.‐Y. Production of multivalent protein binders using a self‐trimerizing collagen‐like peptide scaffold. FASEB J. 22, 3795–3804 (2008)