Potentiation of C1-esterase inhibitor by heparin and interactions with C1s protease as assessed by surface plasmon resonance |
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Authors: | Mohsen RajabiEvi Struble Zhaohua ZhouElena Karnaukhova |
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Institution: | a Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD, 20892 USAb Laboratory of Plasma Derivatives, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD, 20892 USAc Division of Monoclonal Antibodies, Center for Drugs Evaluation and Research, Food and Drug Administration, Bethesda, MD, 20892 USA |
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Abstract: | BackgroundHuman C1-esterase inhibitor (C1-INH) is a multifunctional plasma protein with a wide range of inhibitory and non-inhibitory properties, mainly recognized as a key down-regulator of the complement and contact cascades. The potentiation of C1-INH by heparin and other glycosaminoglycans (GAGs) regulates a broad spectrum of C1-INH activities in vivo both in normal and disease states.Scope of researchWe have studied the potentiation of human C1-INH by heparin using Surface Plasmon Resonance (SPR), circular dichroism (CD) and a functional assay. To advance a SPR for multiple-unit interaction studies of C1-INH we have developed a novel (consecutive double capture) approach exploring different immobilization and layout.Major conclusionsOur SPR experiments conducted in three different design versions showed marked acceleration in C1-INH interactions with complement protease C1s as a result of potentiation of C1-INH by heparin (from 5- to 11-fold increase of the association rate). Far-UV CD studies suggested that heparin binding did not alter C1-INH secondary structure. Functional assay using chromogenic substrate confirmed that heparin does not affect the amidolytic activity of C1s, but does accelerate its consumption due to C1-INH potentiation.General significanceThis is the first report that directly demonstrates a significant acceleration of the C1-INH interactions with C1s due to heparin by using a consecutive double capture SPR approach. The results of this study may be useful for further C-INH therapeutic development, ultimately for the enhancement of current C1-INH replacement therapies. |
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Keywords: | Anti-C1-INH ab anti-C1-esterase inhibitor antibody α1-PI α1-proteinase inhibitor ATIII antithrombin C1-INH C1-esterase inhibitor CD circular dichroism DMSO dimethyl sulfoxide GAG glycosaminoglycan L/P ligand-to-protein molar ratio PBS phosphate buffer saline RCL reactive center loop SA Streptavidin SPR Surface Plasmon Resonance |
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