2-(2-Hydroxy-5-nitrobenzylidene)-1,3-indanedione versus Fluorescein Isothiocyanate in Interaction with Anti-hFABP Immunoglobulin G1: Fluorescence Quenching, Secondary Structure Alteration and Binding Sites Localization

International Journal of Molecular Sciences

Dana Stan¹  Carmen-Marinela Mihailescu³  Mihaela Savin² 

[1] National Institute for R&D in Microtechnologies (IMT), 126A Erou Iancu Nicolae, Bucharest 077190, Romania;Department of Physical Chemistry, Faculty of Chemistry, University of Bucharest, Bd. Regina Elisabeta, No. 4–12, Bucharest 030018, Romania;DDS Diagnostic, Gh. Sincai Street No. 9A, Bucharest 040312, Romania; E-Mail:

关键词: indane-1;    3-dione derivative;    fluorescein isothiocyanate;    immunoglobulin G1;    fluorescence spectroscopy;    circular dichroism;    molecular modeling;   

DOI  :  10.3390/ijms14023011

来源: mdpi

【 摘 要 】

The first step in determining whether a fluorescent dye can be used for antibody labeling consists in collecting data on its physical interaction with the latter. In the present study, the interaction between the 2-(2-hydroxy-5-nitrobenzylidene)-1,3-indanedione (HNBID) dye and the IgG1 monoclonal mouse antibody anti-human heart fatty acid binding protein (anti-hFABP) has been investigated by fluorescence and circular dichroism spectroscopies and complementary structural results were obtained by molecular modeling. We have determined the parameters characterizing this interaction, namely the quenching and binding constants, classes of binding sites, and excited state lifetimes, and we have predicted the localization of HNBID within the Fc region of anti-hFABP. The key glycosidic and amino acid residues in anti-hFABP interacting with HNBID have also been identified. A similar systematic study was undertaken for the well-known fluorescein isothiocyanate fluorophore, for comparison purposes. Our results recommend HNBID as a valuable alternative to fluorescein isothiocyanate for use as a fluorescent probe for IgG1 antibodies.

【 授权许可】

CC BY   
© 2013 by the authors; licensee Molecular Diversity Preservation International, Basel, Switzerland.