Interaction between Chromosomal Protein HMGB1 and DNA Studied by DNA-Melting Analysis

Interaction of HMGB1 nonhistone chromosomal protein with DNA was studied using circular dichroism spectroscopy and thermal denaturation of DNA. Melting DNA in the complex was shown to be a biphasic process. The characteristic melting temperatures of unbound DNA and the DNA bound to HMGB1 in 0.25 mM EDTA solutions were found to be TmI=44.0±0.5°C and TmII=62.0±1°C, respectively. It was shown that the binding of the HMGB1 molecule affects the melting of the DNA region approximately 30 b.p. long

Fluorescent silver clusters on protein templates: understanding their structure

Luminescent metal nanoclusters (NCs) stabilized by natural proteins are of special interest in bioimaging applications. However, the detailed structure of the protein-templated NCs and the nature of their emissive states remain poorly understood. A fair amount of nonluminescent metal ions and clusters complexed to the proteins hinders probing of the structure of the emitting clusters using mass spectroscopy, infrared, or other conventional spectroscopy methods. In this respect, only luminescent excitation spectra distinguish the emitting NCs. In this experimental and theoretical joint study, we modeled the fluorescent excitation and excitation anisotropy spectra of protein-based silver (Ag) NCs. We varied the synthesis conditions and studied the spectral properties of Ag clusters on bovine serum albumin (BSA) and lysozyme, which had already been used as templates, as well as on HMG box (HMGB1) and histone H1 (H1) proteins. We also calculated the electronic spectra of quantum mechanics-optimized Ag–thiolate, Ag–semiquinone, and Ag–formaldehyde complexes with two confined electrons using second-order algebraic diagrammatic construction [ADC(2)] and resolution-of-identity approximate coupled-cluster singles-and-doubles (RI-CC2) methods and compared them with the experimental spectra. We propose a model for the fluorescent Ag–protein complexes in which two reduced Ag atoms are sufficient to form the fluorescent core of the complex. The proposed structural model of the luminescent centers in the Ag–protein complexes differs from the common view that the fluorescent metal NCs in proteins contain about 10 or more metal atoms. The fluorescent Ag clusters formed on the four investigated natural protein matrices exhibited two different spectral and structural patterns. Deprotonated free cysteine residues stabilized the fluorescent Ag3+1 core formed in the BSA matrix. The second type of fluorescent center was realized in the H1, HMGB1, and lysozyme protein matrixes. In this case, tyrosine residues probably stabilize the fluorescent Ag2 centers

Fluorescent silver clusters on protein templates: understanding their structure

Luminescent metal nanoclusters (NCs) stabilized by natural proteins are of special interest in bioimaging applications. However, the detailed structure of the protein-templated NCs and the nature of their emissive states remain poorly understood. A fair amount of nonluminescent metal ions and clusters complexed to the proteins hinders probing of the structure of the emitting clusters using mass spectroscopy, infrared, or other conventional spectroscopy methods. In this respect, only luminescent excitation spectra distinguish the emitting NCs. In this experimental and theoretical joint study, we modeled the fluorescent excitation and excitation anisotropy spectra of protein-based silver (Ag) NCs. We varied the synthesis conditions and studied the spectral properties of Ag clusters on bovine serum albumin (BSA) and lysozyme, which had already been used as templates, as well as on HMG box (HMGB1) and histone H1 (H1) proteins. We also calculated the electronic spectra of quantum mechanics-optimized Ag–thiolate, Ag–semiquinone, and Ag–formaldehyde complexes with two confined electrons using second-order algebraic diagrammatic construction [ADC(2)] and resolution-of-identity approximate coupled-cluster singles-and-doubles (RI-CC2) methods and compared them with the experimental spectra. We propose a model for the fluorescent Ag–protein complexes in which two reduced Ag atoms are sufficient to form the fluorescent core of the complex. The proposed structural model of the luminescent centers in the Ag–protein complexes differs from the common view that the fluorescent metal NCs in proteins contain about 10 or more metal atoms. The fluorescent Ag clusters formed on the four investigated natural protein matrices exhibited two different spectral and structural patterns. Deprotonated free cysteine residues stabilized the fluorescent Ag3+1 core formed in the BSA matrix. The second type of fluorescent center was realized in the H1, HMGB1, and lysozyme protein matrixes. In this case, tyrosine residues probably stabilize the fluorescent Ag2 centers