Kinetic Studies on the Reaction between Dicyanocobinamide and Hypochlorous Acid

Abstract

Hypochlorous acid (HOCl) is a potent oxidant generated by myeloperoxidase (MPO), which is an abundant enzyme used for defense against microbes. We examined the potential role of HOCl in corrin ring destruction and subsequent formation of cyanogen chloride (CNCl) from dicyanocobinamide ((CN)2-Cbi). Stopped-flow analysis revealed that the reaction consists of at least three observable steps, including at least two sequential transient intermediates prior to corrin ring destruction. The first two steps were attributed to sequential replacement of the two cyanide ligands with hypochlorite, while the third step was the destruction of the corrin ring. The formation of (OCl)(CN)-Cbi and its conversion to (OCl)2-Cbi was fitted to a first order rate equation with second order rate constants of 0.002 and 0.0002 µM−1s−1, respectively. The significantly lower rate of the second step compared to the first suggests that the replacement of the first cyanide molecule by hypochlorite causes an alteration in the ligand trans effects changing the affinity and/or accessibility of Co toward hypochlorite. Plots of the apparent rate constants as a function of HOCl concentration for all the three steps were linear with Y-intercepts close to zero, indicating that HOCl binds in an irreversible one-step mechanism. Collectively, these results illustrate functional differences in the corrin ring environments toward binding of diatomic ligands.

Melatonin interferes with COVID-19 at several distinct ROS-related steps

“…Furthermore, an initial rate of 8 × 10 3 M −1 s −1 was found for the binding of HOCl to hemoglobin, further supporting the possibility of HOCl-mediated hemoglobin damage in a biological setting [ 12 , 43 ]. Similarly, an initial rate constant of 2 × 10 3 M −1 s −1 was found for the binding of HOCl to a vitamin B12 derivative [ 8 , 14 ]. In a previous study, 200 μM HOCl was found to react with and destroy the heme of hemoglobin in red blood cells [ 12 ].…”

“…Using a variety of biochemical, physiological, and kinetic techniques, it has been shown that melatonin ceases HOCl-mediated vitamin B12 derivative corrin destruction [ 8 , 14 ]. Vitamin B12 deficiency has been associated with several conditions related to memory loss, immune system disorders, aging, heart disease, male infertility, diabetes, sleep disorders, depression, mental disorders, and inflammation [ 71 ].…”

“…1 ). HOCl exists in approximate equilibrium with hypochlorite ion ( − OCl) at normal body pH 7.4 (pKa 7.59); however only the uncharged molecule can easily penetrate the cell membrane, most likely by passive diffusion through porins, proteins on the cell membrane that form channels [ 8 , 9 ]. MPO can be attached to extracellular NETs, be released at the plasma membrane due to inappropriate trafficking control, bind to the glycocalyx of several cell types and further increase neutrophil recruitment, or otherwise leak from the phagolysosome.…”

“…All of these events can cause additional catalysis of HOCl [ 10 ]. In addition, the production of excessive amounts of ROS can interfere with the catalytic site of many hemoprotein model compounds, ultimately contributing to malfunction and/or destruction of hemoglobin and red blood cells, nitric oxide synthase (NOS), catalase, corrin rings such as in vitamin B12, zinc-Cys clusters, and sulfur‑sulfur bonds, deficiencies of which are characteristic of COVID-19 [ 8 , [11] , [12] , [13] , [14] , [15] , [16] ]. HOCl might also modulate other important pathological mechanisms in COVID-19 including the release of free iron (Fe(II)) from HOCl mediated hemoprotein destruction, which reacts with HOCl directly or through the Fenton reaction generating • OH.…”

Melatonin prevents hypochlorous acid‐mediated cyanocobalamin destruction and cyanogen chloride generation

“…Recently, utilizing rapid kinetic measurements, high‐performance liquid chromatography (HPLC), and mass spectrometric techniques, we have highlighted the mechanism through which HOCl mediates the destruction of the metal‐ion derivatives of tetrapyrrole macrocyclic rings such as hemoproteins and vitamin B 12 (VB 12 ) derivatives . We have shown destruction of cyanocobalamin (Figure A), a common pharmacological form of VB 12 , mediated by HOCl, resulted in the generation of toxic molecular products such as chlorinated derivatives, different degradation products of corrin, the organic ring system of cobalamin, release of the toxic blood agents (CN − /cyanogen chloride [CNCl]), and redox‐active free cobalt (Co) through the formation of chloro‐cyanocobalamin (Figure B) …”