Thermodynamic Stability and Functional Activity of Tumor-Associated Antibodies

O. P. Bliznukov¹, L. D. Kozmin¹, V. B. Klimovich², Y. I. Tsybovsky³, Z. I. Kravchuk³, and S. P. Martsev^3*

¹St. Petersburg Institute for Nuclear Physics, Russian Academy of Sciences, Orlova Roshcha, Gatchina, 188300 Russia; fax: (81271) 32-302; E-mail: leokon@yahoo.com

²Central Research Roentgeno-Radiological Institute, Poselok Pesochnoe, St. Petersburg, 188646 Russia

³Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, ul. Akademika Kuprevicha 5/2, Minsk, 220141 Belarus; fax: +375-17-2-63-72-74; E-mail: martsev@ns.iboch.ac.by

^* To whom correspondence should be addressed.

Received July 5, 2000

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Tumor-associated antibodies of human IgG1 subclass were eluted from cell-surface antigens of human carcinoma cells and studied by differential scanning calorimetry and binding to local conformational probes, protein A from Staphylococcus aureus and a monoclonal antibody targeted to the CH2 domain of the Fc fragment. At pH 2.0-7.0, we observed virtually identical enthalpies of thermal unfolding for IgG1 from normal human sera and tumor-associated IgG1. The exact values of calorimetric enthalpy (Deltah) at pH 7.0 were 6.1 and 6.2-6.3 cal/g for IgG1 from normal serum and IgG1 from carcinoma cells, respectively. The affinity constants of protein A binding to the CH2-CH3 domain interface demonstrated differences between serum IgG1 and tumor associated IgG1 that did not exceed 3-8-fold. The binding affinity toward the anti-CH2 monoclonal antibody determined for serum IgG1 and IgG1 from carcinoma cells differed not more than 2.5-fold. The thermodynamic parameters of IgG1 from carcinoma cells strongly suggest that protein conformational stability was essentially unaltered and that the Fc fragment of the tumor-derived IgG1 preserved its structural integrity.

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KEY WORDS: IgG1, carcinoma, antitumor antibodies, differential scanning calorimetry, conformational stability, protein A

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