Highly specific inhibition of C1q globular-head binding to human IgG: a novel approach to control and regulate the classical complement pathway using an engineered single chain antibody variable fragment.
We sought to specifically regulate the binding of human C1q, and thus the activation of the first complement component, via the construction of a single chain antibody variable binding region fragment (scFv) targeting the C1q globular heads. Here we describe details of the construction, expression and evaluation of this scFv, which was derived from a high-affinity hybridoma (Qu) specific for the C1q globular heads. The scFv was comprised of the Qu variable heavy chain domain (VH) linked to the Qu variable light chain domain (VL) and was termed scFv-QuVHVL. When mixed with either purified C1q or with human serum as a source of C1, scFv-QuVHVL bound to C1q and competitively restricted the interaction of C1q or C1 with immobilized IgG or with IgG1 antibody-coated cells, and prevented the activation of native C1 in human serum as determined by analyses of C1-mediated C4 deposition and fluid-phase C4 conversion. However scFv-QuVHVL could be manipulated to become a C1 activator when it was irreversibly immobilized onto microtiter ELISA plates, prior to contact with human serum complement. This functional dichotomy can be a useful tool in selectively elucidating, differentiating, inducing or inhibiting specific roles of human C1q and the classical complement pathway in complement-mediated physiological processes. We project that once fully humanized, fluid-phase scFv-QuVHVL could become a useful therapeutic in limiting inadvertent host tissue damage elicited by the classical complement pathway.
Hwang
Hee Young
HY
Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, SC 29464, USA.
Duvall
Marcus R
MR
Tomlinson
Stephen
S
Boackle
Robert J
RJ
D015922:Complement C1q / Q000276:immunology / Q000378:metabolism*; D003171:Complement Pathway, Classical; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism*; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000378:metabolism*; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D015922:Complement C1q