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Characterization of the Interactions of Lysozyme with DNA by Surface Plasmon Resonance and Circular Dichroism Spectroscopy

Author:
Lin, Kuo-Chih, Wey, Ming-Tsai, Kan, Lou-Sing, Shiuan, David
Source:
Applied biochemistry and biotechnology 2009 v.158 no.3 pp. 631-641
ISSN:
0273-2289
Subject:
DNA, DNA replication, DNA-binding domains, Human immunodeficiency virus 1, circular dichroism spectroscopy, dissociation, electrostatic interactions, gene expression, immunity, lysozyme, nucleotide sequences, peptides, surface plasmon resonance
Abstract:
Association with nucleic acid has been recognized as a unique role of lysozyme and may explain why lysozyme was called a killer protein against HIV infection. In the present study, we characterized the interactions of lysozyme and its derived peptides with a biotin-labeled pUC19 plasmid DNA. Real-time detection of the macromolecular interaction was performed using the SPR (surface plasmon resonance) spectroscopy. The SPR sensorgrams were analyzed and the association and dissociation rate constants as well as the dissociation equilibrium constant K D were, thus, estimated. The results reveal that other than the electrostatic interactions between the basic protein and the nucleotide sequences carrying negative charges, the specific DNA-binding motifs at the N- and C-termini of lysozyme were also involved in the interactions. The nonapeptide RAWVAWRNR (aa 107-115 of lysozyme) reported previously to block HIV-1 viral entrance and replication was also able to bind DNA with its K D value comparable to that of histones. The possibilities of ligand-binding-induced conformational changes were investigated using the circular dichroism spectroscopy. The CD spectra (200-320 nm) reveal that the conformational changes indeed occur as the spectra of lysozyme-DNA interactions are much less at the major trough region than the sum of individual spectra. The interaction of lysozyme with DNA molecules may interfere with DNA replication, modulate gene expression, and block bacterial and viral infections. These all suggest that human lysozyme may represent part of the innate immune system with a very broad protective spectrum.
Agid:
638681