Oxidative degradation of biological substrates by hypochlorous acid has been examined under reaction conditions similar to those found in active phagosomes. Iron sulfur proteins are bleached extremely rapidly, followed in decreasing order by f3carotene, nucleotides, porphyrins, and heme proteins. Enzymes containing essential cysteine molecules are inactivated with an effectiveness that roughly parallels the nucleophilic reactivities of their sulfhydryl groups. Other compounds, including glucosamines, qumiones, riboflavin, and, except for N-chlorination, phospholipids, are unreactive. Rapid irreversible oxidation of cytochromes, adenine nucleotides, and carotene pigments occurs when bacterial cells are exposed to exogenous hypochlorous acid; with Escherichia coli, titrimetric oxidation of cytochrome was found to coincide with loss of aerobic respiration. The occurrence of these cellular reactions implicates hypochlorous acid as a primary microbicide in myelo.roxidase-containing leukocytes; the reactivity patterns observed are consistent with the view that bactericidal action results primarily from loss of energy-linked respiration due to destruction of cellular electron transport chains and the adenine nucleotide pool.Bacteria commonly lose their ability to divide within minutes of encountering phagocytosing leukocytes (1). Loss ofcell viability often occurs well before the onset ofcellular digestion as determined by physiological changes (2), macromolecular degradation (3, 4), or loss of macromolecular biosynthesis (5). The specific reactions giving rise to cellular death have not yet been identified, at least in part because leukocytes are capable ofinitiating a diverse set of processes which are potentially lethal (6, 7). Given the above observations, however, the microbicidal reactions must be among the first that attend interaction with the leukocyte.Reactions catalyzed by myeloperoxidase (MPOase) appear to make major contributions to the microbicidal-action of polymorphonuclear leukocytes (PMNs) (6, 7). The cell-free MPOase-H202-Cl-system is 'potentially microbicidal; chlorination of bacteria by the cell-free system (8, 9) and of macromolecular fractions during PMN digestion of bacteria (9) electrophile-nucleophile interactions involving association of electropositive chlorine with electron-rich centers on the substrate; reaction pathways are-correspondingly highly dependent upon medium-conditions, particularly H' and Cl-concentrations (14). With the exception ofamines and amino acids (11,12,15), the reactions of biological compounds with HOC1 are not understood (16).We report 'here the results of a survey of simple biological compounds which can be taken as prototypic ofvarious components of bacterial cells. :The data demonstrate that HOCI is strongly selective -toward nucleotides and compounds that are models for certain components ofrespiratory redox chains. This selectivity is shownito extend to bacterial cells. Table 1 were -determined from loss of their characteristic absorption bands when ...

Titrimetric addition of hypochlorous acid (HOCI) or chloramine (NH2Cl) to suspensions of Escherichia coli decreases their ability to accumulate "C-labeled glutamine, proline, thiomethylgalactoside, and leucine in a manner that approximately coincides with loss of cell viability; quantitative differences in cellular response are observed with the two oxidants. Inhibition of j-galactosidase activity in E. coli a

No abstract

No abstract

Oxidation of Escherichia coli by hypochlorous acid (HOCl) or chloramine (NH2Cl) gives rise to massive hydrolysis of cytosolic nucleotide phosphoanhydride bonds, although no immediate change occurs in either the nucleotide pool size or the concentrations of extracellular end products of AMP catabolism. Titrimetric curves of the extent of hydrolysis coincide with curves for loss of cell viability, e.g., reduction in the adenylate energy charge from 0.8 to 0.1-0.2 accompanies loss of 99% of the bacterial CFU. The oxidative damage caused by HOCl is irreversible within 100 ms of exposure of the organism, although nucleotide phosphate bond hydrolysis requires several minutes to reach completion. Neither HOCl nor NH2Cl reacts directly with nucleotides to hydrolyze phosphoanhydride bonds. Loss of viability is also accompanied by inhibition of induction of beta-galactosidase. The proton motive force, determined from the distribution of 14C-radiolabeled lipophilic ions, declines with incremental addition of HOCl after loss of respiratory function; severalfold more oxidant is required for the dissipation of the proton motive force than for loss of viability. These observations establish a causal link between loss of metabolic energy and cellular death and indicate that the mechanisms of oxidant-induced nucleotide phosphate bond hydrolysis are indirect and that they probably involve damage to the energy-transducing and transport proteins located in the bacterial plasma membrane.