Oxidation of hydroquinone by myeloperoxidase. Mechanism of stimulation by benzoquinone.

Myeloperoxidase (MPO) is a prime candidate for mediating the inflammatory tissue damage of neutrophils because it converts Cl- to the potent oxidant hypochlorous acid. It also oxidizes xenobiotics to reactive free radicals. We have found that the kinetics of oxidation of hydroquinone by myeloperoxidase are inadequately explained by the classical peroxidase mechanism. Peroxidation of hydroquinone displayed a distinct lag phase, which was practically abolished by excluding O2 and was eliminated by adding benzoquinone at the start of the reaction. Superoxide dismutase increased the rate of peroxidation by 40% but did not eliminate the lag phase. Spectral investigations revealed that during the initial phase of the reaction, MPO was converted to oxy-MPO, or compound III, by a mechanism that was not reliant on superoxide. Benzosemiquinone, however, was able to convert ferric-MPO to compound III. Both compound III and ferro-MPO reacted with benzoquinone to regenerate ferric-MPO. We propose that the lag phase occurs because benzosemiquinone reduces ferric-MPO to ferro-MPO, which rapidly binds O2 to form compound III. Since compound III is outside the peroxidation cycle, conversion of hydroquinone to benzoquinone is retarded. However, as benzoquinone accumulates, it oxidizes ferro-MPO and compound III to ferric-MPO, thereby increasing the rate of peroxidation. There is a minimal lag phase under an atmosphere of N2 because ferro-MPO would be rapidly oxidized by benzoquinone, without formation of compound III. We conclude that when substrates produce radicals capable of reducing ferric-MPO, they will be peroxidized efficiently only if oxy-MPO is readily recycled. Furthermore, these radicals will prevent MP3+ from reacting with H2O2, and thereby prevent the enzyme from oxidizing Cl- to hypochlorous acid. Thus, this mechanism could be exploited to prevent hypochlorous acid-mediated inflammatory tissue damage.     

Inhibition of Babesia spp. by plastics.

Five commercially available plastic containers were compared with glass for toxicity toward Babesia rodhaini and Babesia bigemina. Comparisons were made by using infectivity tests in mice (B. rodhaini) and cattle (B. bigemina). Low-density polypropylene and polystyrene containers were not toxic, but two of the three polyvinyl chloride containers tested significantly reduced the viability of both species of Babesia. Plasticizer present in various amounts on the surface of the toxic containers was most likely the inhibitory material.     

Protein modifications by activated carcinogens. II. The acetylation of ribonuclease by N-acetoxy-3-fluorenylacetamide.

The reaction of N-[3H]acetoxy-3-fluorenylacetamide (N-[3H]acetoxy-3-FAA), a potent carcinogen for the rat, with RNAase yielded three modified proteins separable from RNAase by ion exchange chromatography on Bio-Gel CM-30 with a gradient of increasing sodium ion concentration. Only minor amounts of RNAase were recovered. The modified proteins were labeled with 3H to a varying degree, and their order of elution was inversely related to the extent of labeling. The modification of the proteins was the result of the transfer of the acetyl group from N-[3H]acetoxy-3-FAA to RNAase. The evidence for this conclusion was (a) the release of 84-86% of the radioactivity as [3H] acetic acid from the two major proteins upon acid hydrolysis and (b) the isolation of eplision-N-[3H] acetyl-L-lysine from enzymatic hydrolysates of these proteins. A comparison of the present data with those previously reported for the acetylaton of RNAase by the isomeric carcinogen, N-acetoxy-2-FAA, showed that N-acetoxy-3-FAA is the more potent acetyl-lating agent. The present study in conjunction with the previous results, suggests that structural alteration of cellular nucleopholes by acylation may be a biochemical mechanism underlying the biological activity of N-acetoxy-3-FAA and related activated carcinogens.     

Production of lymphotoxin by human lymphocytes. II. Stimulation by purified tuberculin]

A total of 43 lymphocyte culture (LC) from 12 adults (aged 32--48 years) and 21 children 6--7 years of age were studied. In PPD-stimulated adults LC stimulation ratio ranged from 8 to 39%; their cytotoxic activity, i.e. lymphotoxin (LT) production, was determined by staining target L cells with crystal violet and measuring protein synthesis in surviving target cells by radioactive amino acid incorporation. In children blastogenic response occurred in 11 out of 21 PPD-stimulated LC studied, but only 8 supernatant culture fluids were toxic to L cells and 3 were not. These findings were confirmed by repeated tests 3 weeks later. Probable correlation of BTL and LT production with the functional state of specific immunity to tuberculosis is discussed.     

Inhibition of tyrosinase by indole compounds and reaction products. Protection by albumin.

Tyrosinase activity decreases as the reaction proceeds and is inhibited by L-3,4-dihydroxyphenylalanine oxidation products. Indole and tryptophan inhibit tyrosinase reaction and bovine albumin protects against end-product(s) inhibition or inactivation. Since the same tyrosinase reaction products are indole compounds and some authors reported the binding of indole derivatives with albumin, it is here suggested that indole intermediates of melanin synthesis inhibit or inactivate tyrosinase.