| Autor: |
Gorazd Vesnaver, Črtomir Podlipnik, Jurij Lah, Drago Kuzman, Simona Jevševar, Jaka Marušič |
| Rok vydání: |
2012 |
| Předmět: |
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| Zdroj: |
Journal of Biological Chemistry. 287:8613-8620 |
| ISSN: |
0021-9258 |
| DOI: |
10.1074/jbc.m111.318451 |
| Popis: |
Human tumor necrosis factor α (TNF-α) exists in its functional state as a homotrimeric protein and is involved in inflammation processes and immune response of a human organism. Overproduction of TNF-α results in the development of chronic autoimmune diseases that can be successfully treated by inhibitors such as monoclonal antibodies. However, the nature of antibody-TNF-α recognition remains elusive due to insufficient understanding of its molecular driving forces. Therefore, we studied the energetics of binding of a therapeutic antibody fragment (Fab) to the native and non-native forms of TNF-α by employing calorimetric and spectroscopic methods. Global thermodynamic analysis of data obtained from the corresponding binding and urea-induced denaturation experiments has been supported by structural modeling. We demonstrate that the observed high affinity binding of Fab to TNF-α is an enthalpy-driven process due mainly to specific noncovalent interactions taking place at the TNF-α-Fab binding interface. It is coupled to entropically unfavorable conformational changes and accompanied by entropically favorable solvation contributions. Moreover, the three-state model analysis of TNF-α unfolding shows that at physiological concentrations, TNF-α may exist not only as a biologically active trimer but also as an inactive monomer. It further suggests that even small changes of TNF-α concentration could have a considerable effect on the TNF-α activity. We believe that this study sets the energetic basis for understanding of TNF-α inhibition by antibodies and its unfolding linked with the concentration-dependent activity regulation. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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