Journal
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
Volume 793, Issue 1, Pages 93-105Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/S1570-0232(03)00367-2
Keywords
squarylium dyes; non-covalent protein labeling
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Noncovalent interactions between two squarylium dyes and various model proteins have been explored. NN 127 and SQ-3 are symmetric and asymmetric squarylium dyes, respectively, the fluorescence emissions of which have been shown to be enhanced upon complexation with proteins such as bovine serum albumin (BSA), human serum albumin (HSA), P-lactoglobulin A, and trypsinogen. Although these dyes are poorly soluble in aqueous solution, they can be dissolved first in methanol followed by dilution with aqueous buffer without precipitation, and are then suitable for use as fluorescent labels in protein determination studies. The nature of interactions between these dyes and proteins was studied using a variety of buffer systems, and it was found that electrostatic interactions are involved but not dominant. Dye/protein stoichiometries in the noncovalent complexes were found to be 1: 1 for SQ-3, although various possible stoichiometries were found for NN 127 depending upon pH and protein. Association constants on the order of 10(5) and 10(7) were found for noncovalent complexes of SQ-3 and NN127, respectively, with HSA, indicating stronger interactions of the symmetric dye with proteins. Finally, HSA complexes with NN 127 were determined by capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). In particular, NN127 shows promise as a reagent capable of fluorescently labeling analyte proteins for analysis by CE-LIF without itself being significantly fluorescent under the aqueous solution conditions studied herein. (C) 2003 Elsevier B.V. All rights reserved.
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