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 Title: Effects of Secondary Forces on the Primary Interaction and Variable Domain Conformation of Antifluorescein Antibodies Author(s): Mummert, Mark Eric Doctoral Committee Chair(s): Voss, Edward W., Jr. Department / Program: Microbiology Discipline: Microbiology Degree Granting Institution: University of Illinois at Urbana-Champaign Degree: Ph.D. Genre: Dissertation Subject(s): Health Sciences, Immunology Abstract: Secondary forces are biochemical interactions that occur between regions surrounding the mouth of the antibody active site and the environment of homologous ligand or epitope. These interactions have been demonstrated to modulate the antibody variable domains as well as impacting complex stability. Although secondary forces do not dictate antibody specificity (the primary interaction) an understanding of these interactions is critical in evaluating the nuances of antibody/antigen interactions. Described within this thesis is a series of biophysical measurements that have allowed the delineation and quantitation of secondary forces. These are the first experiments to convincingly separate primary interactive components from secondary interactive components. This dissection of primary from secondary forces was made possible by the utilization of the monoclonal antifluorescein antibody system. Fluorescein has been shown to be an active site-filling moiety. Fluorescein 5-isothiocyanate was covalently linked to the $\varepsilon$-amine of lysine residues in proteins or synthetic peptides. Differences in the binding reactivity and various spectral parameters of a series of monoclonal antifluorescein antibodies were compared and contrasted when bound with fluorescein and fluorescein conjugated to unique physical and chemical topolgies. Since the active site was completely filled by the fluorescyl moiety, differences in measured properties was a consequence of attached residues that composed the carrier environment. Three monoclonal antifluorescein antibodies (mAbs) were utilized for these studies (4-4-20, 9-40 and 18-2-3). This panel of antibodies was chosen due to the high degree with which they have been characterized biochemically and biophysically. Differential effects due to secondary forces were correlated with known properties of these antibodies. Moreover, these antibodies were compared and contrasted with one another. Models employing variable domain dynamics were developed to explain results from the various experiments and were correlated with known structural and chemical information regarding these antibodies. Issue Date: 1997 Type: Text Language: English Description: 234 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1997. URI: http://hdl.handle.net/2142/86719 Other Identifier(s): (MiAaPQ)AAI9737206 Date Available in IDEALS: 2015-09-28 Date Deposited: 1997
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