کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2188072 1096151 2007 15 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Comparative Analyses of Complex Formation and Binding Sites between Human Tumor Necrosis Factor-alpha and its Three Antagonists Elucidate their Different Neutralizing Mechanisms
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیولوژی سلول
پیش نمایش صفحه اول مقاله
Comparative Analyses of Complex Formation and Binding Sites between Human Tumor Necrosis Factor-alpha and its Three Antagonists Elucidate their Different Neutralizing Mechanisms
چکیده انگلیسی

Tumor necrosis factor-alpha (TNFα)-blocking therapy, using biologic TNFα antagonists, has been approved for the treatment of several diseases including rheumatoid arthritis, psoriasis and Crohn's disease. There have been few detailed studies of binding characterizations for the complex formation by TNFα and clinically relevant antagonists, particularly Infliximab (Remicade®) and Etanercept (Enbrel®). Here we characterized the binding stoichiometry and size of soluble TNFα–antagonist complexes and identified energetically important binding sites on TNFα for the three antagonists, Etanercept, Infliximab, and the recently developed humanized TNFα neutralizing monoclonal antibody, YHB1411-2. Size-exclusion chromatography and dynamic light scattering analyses revealed that the three antagonists formed distinct thermodynamically stable TNFα–antagonist complexes that exhibited differences in their size and composition. Energetically important binding residues on TNFα were identified for each antagonist by a sequence of experiments that consisted of competition binding assays, fragmentations, loop mutations, and single-point mutations using yeast surface-displayed TNFα, which was further confirmed for solubly purified TNFα mutants by surface plasmon resonance technique. Analyses of the binding geometry based on binding site location, spatial constraints, and valency satisfaction allowed us to interpret the thermodynamically stable complexes as follows: one molecule of Etanercept and one molecule of trimeric TNFα (Etanercept1––TNFα1), Infliximab6–TNFα3, and YHB1411–24-TNFα2. The distinct features of the soluble antagonist–TNFα complex formation among the antagonists may give further insights into their different neutralizing mechanisms and pharmacokinetic profiles.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Molecular Biology - Volume 374, Issue 5, 14 December 2007, Pages 1374–1388
نویسندگان
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