No abstract

1-Phosphohistidine and 3-phosphohistidine Phosphohistidine has been isolated as the solid lithium have been synthesized by reaction of histidine with salt and has been characterized by composition, ultraphosphoramidate. The two isomers have been sepaviolet and nuclear magnetic resonance spectra, titrarated by paper electrophoresis and anion-exchange tion curve, and rate of hydrolysis over a wide range of chromatography. The synthetic phosphohistidine isopH values. a-N-Acetyl-3-phosphohistidine, l-methylmer which is identical with the compound isolated from 3-phosphohistidine, a-AT-acetyl-1-methyl-3-phosphomitochondria and from a succinate thiokinase preparahistidine, and the isomers of phosphohistidine methyl tion has been identified as 3-phosphohistidine. 3-ester have been prepared and some properties reported.

I. Phosphorylated derivatives of histidine were synthesized and studied in order to provide a better understanding of the chemistry of phosphohistidine-containing proteins, i-Phosphohistidine, 1,3-diphosphohistidine, a, 1,3-triphosphohistidine, and phosphorylated derivatives of ~-N-acetylhistidine were prepared by reactions with POC13 or phosphoramidate. The compounds were purified by ion-exchange chromatography and characterized by the phosphate:histidine ratio, electrophoretic behavior, and ultraviolet spectra. 2. The rates of hydrolysis of these compounds, as well as 3-phosphohistidine, phosphoimidazole, and 1,3-diphosphoimidazole, were determined over a wide range of H + concentrations. In strongly acidic solution all of the compounds appeared to be hydrolyzed by the same mechanism. In all cases the N-phosphoryl bond was hydrolyzed more readily if a proton or phosphoryl group were substituted on the other ring nitrogen, and histidine derivatives were less stable than the corresponding imidazole derivatives. 3. For both mono-and diphosphoryl derivatives of histidine, the N-I phosphoryl group was hydrolyzed and transferred to other imidazole compounds more readily than the N-3 phosphoryl group. The greater reactivity at the N-I position is partially due to the presence of the protonated a-amino group. 4. In aqueous solution, I-phosphohistidine is rapidly converted to 3-phosphohistidine, histidine and inorganic phosphate.

I. An enzyme that catalyzes the reduction of erythrocyte cytochrome b 5 has been isolated from the supernatant fraction of erythrocyte hemolysates by chromatography on DEAE-cellulose, Amberlite CG-5 o, and Bio-Gel P-6o. 2. Erythrocyte cytochrome b 5 reductase has been shown to contain FAD. Incubation of the reductase in the absence of EDTA results in both the appearance of flavin fluorescence and the loss of enzymatic activity with time. 3. The reductase catalyzes the reduction of erythrocyte cytochrome b 5 5o times faster with NADH than with NADPH. The apparent Km of NADH was calculated to be 1.6. IO-~ M and the turnover number is 128o moles of cytochrome b 5 per min per mole of flavin. The reduction of electron acceptors proceeded in the following decreasing order of rate: K~Fe(CN)6, 2,6-dichlorophenolindophenol (DCIP), cytochrome b 5, methylene blue, cytochrome c, 03, oxidized glutathione, and methemoglobin. 4-The FAD prosthetic group, the substrate specificity, and the effect of ionic strength, pH, and EDTA on activity suggest that the reductase is the same enzyme as NADH dehydrogenase I, the enzyme lacking in many cases of congenital methemoglobinemia. The properties of the reductase, including its molecular weight, are very similar to those of the cytochrome b 5 reductases solubilized from microsomes and mitochondria of other tissues.

These data indicate that riboflavin improves early I/R injury and reduces the development of CAV, most likely due to alloantigen-independent effects such as reduced early graft oxidant stress. Riboflavin administered in the setting of cardiac allograft transplantation appears to be a powerful means to reduce early graft lipid peroxidation, leukocytic infiltration, and cytokine production as well as to suppress the late development of cardiac allograft vasculopathy.