title: Kinetic method for assaying the halogenating activity of myeloperoxidase based on reaction of celestine blue B with taurine halogenamines.
abstract: Myeloperoxidase (MPO) is a challenging molecular target which, if put under control, may allow regulating the development of inflammatory reactions associated with oxidative/halogenative stress. In this paper, a new kinetic method for assaying the halogenating activity of MPO is described. The method is based on measuring the rate of iodide-catalyzed oxidation of celestine blue B (CB) by oxygen and taurine N-chloramine (bromamine). The latter is produced in a reaction of taurine with HOCl (HOBr). CB is not a substrate for the peroxidase activity of MPO and does not react with hydrogen peroxide and superoxide anion radical. Taurine N-chloramine (bromamine) reacts with CB in molar ratio of 1:2. Using the new method, we studied the dependence of MPO activity on concentration of substrates and inhibitors. The specificity of MPO inhibition by non-proteolyzed ceruloplasmin is characterized. The inhibition of taurine N-chloramine production by neutrophils and HL-60 cells in the presence of MPO-affecting substances is demonstrated. The new method allows determining the kinetic parameters of MPO halogenating activity and studying its inhibition by various substances, as well as screening for potential inhibitors of the enzyme.
authors: Department of Molecular Genetics, Research Institute of Experimental Medicine , Saint-Petersburg , Russia.
authors: A V
authors: AV
authors: Sokolov
authors: V A
authors: VA
authors: Kostevich
authors: S O
authors: SO
authors: Kozlov
authors: I S
authors: IS
authors: Donskyi
authors: I I
authors: II
authors: Vlasova
authors: A O
authors: AO
authors: Rudenko
authors: E T
authors: ET
authors: Zakharova
authors: V B
authors: VB
authors: Vasilyev
authors: O M
authors: OM
authors: Panasenko
mesh_terms: D001965:Bromides / Q000737:chemistry; D002570:Ceruloplasmin / Q000378:metabolism; D004396:Coloring Agents / Q000737:chemistry*; D057075:Enzyme Assays / Q000379:methods*; D004791:Enzyme Inhibitors / Q000378:metabolism; D018922:HL-60 Cells; D054879:Halogenation; D006801:Humans; D007700:Kinetics; D009504:Neutrophils / Q000201:enzymology; D010078:Oxazines / Q000737:chemistry*; D009195:Peroxidase / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013654:Taurine / Q000031:analogs & derivatives* / Q000737:chemistry
chemical_list: D001965:Bromides; D004396:Coloring Agents; D004791:Enzyme Inhibitors; D010078:Oxazines; C032714:bromamine; D013654:Taurine; C043410:N-chlorotaurine; D009195:Peroxidase; D002570:Ceruloplasmin; C007063:Celestine Blue
keywords: HOCl; celestine blue B; ceruloplasmin; halogenating activity; myeloperoxidase; taurine