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

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 KD 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 KD 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.

Lysozyme-Induced Transcriptional Regulation of TNF-α Pathway Genes in Cells of the Monocyte Lineage

“…Besides its direct antibacterial enzymatic activity, lysozyme displays other promising biological properties, which may enable its therapeutic use as an immune-modulating agent in the near future. For example, lysozyme has remarkable nucleic acid-binding properties, which may explain its antiviral activity toward HIV-1 [11,12,13]. Moreover, the ability to stimulate the recovery of lymphocyte CD4+/CD8+ ratio in patients in remission after chemotherapy for malignant diseases [14], and to beneficially act in the context of human ageing [15], further highlights the multifaceted immune-modulating properties of this natural product.…”

Protection of blue shrimp (Litopenaeus stylirostris) against the White Spot Syndrome Virus (WSSV) when injected with shrimp lysozyme

“…Early investigation of this protein found that its positive charge rather than its enzymatic activity accounts for its antiviral action [21]. Lysozyme and lysozyme-derived peptides exhibit DNA-binding activity, and previous researchers have suggested that the interaction between lysozyme and DNA molecules may interfere with DNA replication, modulate gene expression, and block bacterial and viral infections, contributing to the proteins' innate immune modulating activity [22]. These proteins are also known to be opsonic in nature and to have the ability to activate the complement system and increase phagocytic activity.…”

Inhibition of nuclear factor kappaB proteins-platinated DNA interactions correlates with cytotoxic effectiveness of the platinum complexes

“…SPR spectroscopy makes possible real-time detection and the determination of binding kinetics and affinities of biomacromolecular interactions2728.…”