Denaturation of Protein by Chlorine Dioxide:  Oxidative Modification of Tryptophan and Tyrosine Residues

Abstract

Oxychlorine compounds, such as hypochlorous acid (HOCl) and chlorine dioxide (ClO2), have potent antimicrobial activity. Although the biochemical mechanism of the antimicrobial activity of HOCl has been extensively investigated, little is known about that of ClO2. Using bovine serum albumin and glucose-6-phosphate dehydrogenase of Saccharomyces cerevisiae as model proteins, here I demonstrate that the antimicrobial activity of ClO2 is attributable primarily to its protein-denaturing activity. By solubility analysis, circular dichroism spectroscopy, differential scanning calorimetry, and measurement of enzymatic activity, I demonstrate that protein is rapidly denatured by ClO2 with a concomitant decrease in the concentration of ClO2 in the reaction mixture. Circular dichroism spectra of the ClO2-treated proteins show a change in ellipticity at 220 nm, indicating a decrease in alpha-helical content. Differential scanning calorimetry shows that transition temperature and endothermic transition enthalpy of heat-induced unfolding decrease in the ClO2-treated protein. The enzymatic activity of glucose-6-phosphate dehydrogenase decreases to 10% within 15 s of treatment with 10 microM ClO2. Elemental analyses show that oxygen, but not chlorine, atoms are incorporated in the ClO2-treated protein, providing direct evidence that protein is oxidized by ClO2. Furthermore, mass spectrometry and nuclear magnetic resonance spectroscopy show that tryptophan residues become N-formylkynurenine and tyrosine residues become 3,4-dihydroxyphenylalanine (DOPA) or 2,4,5-trihydroxyphenylalanine (TOPA) in the ClO2-treated proteins. Taking these results together, I conclude that microbes are inactivated by ClO2 owing to denaturation of constituent proteins critical to their integrity and/or function, and that this denaturation is caused primarily by covalent oxidative modification of their tryptophan and tyrosine residues.

Systematic Evaluation of the Efficacy of Chlorine Dioxide in Decontamination of Building Interior Surfaces Contaminated with Anthrax Spores

“…The antimicrobial activity of CD gas is due to protein denaturation, some of which may be important for spore viability and/or integrity (12,16,24). Recently, Ogata and Shibata (17) demonstrated model peptide denaturation (derived from influenza A virus) via modification of tyrosine and tryptophan residues.…”

Chlorine Dioxide is a Better Disinfectant than Sodium Hypochlorite against Multi-Drug Resistant <i>Staphylococcus aureus</i>, <i>Pseudomonas aeruginosa</i>, and <i>Acinetobacter baumannii</i>

“…In addition, NaClO is easily inactivated in the presence of biological materials such as blood cells and plasma proteins. In comparison, chlorine dioxide (ClO 2 ) is a water-soluble and yellow gas with a strong oxidizing activity (5,6). Earlier studies have observed that ClO 2 has a potent antimicrobial activity against bacteria, fungi, protozoa, and viruses (7)(8)(9)(10)(11).…”

Efficiency of Traditional Water Treatment Plant and Compact Units in Removing Viruses

“…Although the disinfecting power of chlorine is well documented in the literature against all types of microbes including viruses, the detection of Rotaviruses in the drinking water sample in Shark El-Mansoura WTP suggested a failure in the chlorination process which needs attention. Chlorination was reported denature the proteins and causes breakage in the nucleic acid molecules (Ogata, 2007). These should be sufficient to remove all forms of microbes and viruses.…”