The carbonyl reactivity of 3-bromopyruvate and related compounds

The covalent hydration of β-halopyruvic acids and β-halopyruvamides has been investigated in the uv region by stopped-flow techniques. In aqueous solutions of β-bromopyruvate and related compounds, the geminal diol is the predominant form at all pH values. Kinetic investigations have also been carried out on these hydrations. General bases enhance the rate of approaching the equilibrium. Specific acid catalysis was not detected. In the water-catalyzed reaction of pyruvamides, the relationship between the equilibrium constants K and the rate constants of the forward reaction shows that the transition state is more productlike.     

Endogenous phosphorylation of soluble enzymes in human red cells. Cyclic 3',5'-AMP-dependent phosphorylation of phosphofructokinase without detectable regulatory effect

ATP-depleted human red cells have been incubated in a glucose-containing medium with [32P]orthophosphate in the presence and in the absence of cyclic 3',5'-AMP and dibutyril cyclic 3',5'-AMP. Spectrin, pyruvate kinase, phosphofructokinase, glucose-6-phosphate dehydrogenase and hemoglobin A1 have been purified and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein-bound radioactivity has been measured from the sodium dodecyl sulfate polyacrylamide gels and the trichloroacetic acid-precipitated proteins. In the cytosol, the most intense phosphorylation was found for pyruvate kinase whose, in the presence of cyclic AMP, specific radioactivity was comparable to that of the membrane protein and spectrin. In the absence of cyclic nucleotides it was five times less phosphorylated. Phosphofructokinase was only phosphorylated when the red cells were incubated with cyclic nucleotides; the extent of phosphorylation was four times less than for pyruvate kinase. Hemoglobin, glucose-6-phosphate dehydrogenase and a contaminant protein copurified with phosphofructokinase were not phosphorylated: the 'background' of the radioactivity found for these proteins was 100 times less than for pyruvate kinase and spectrin, and 20 times less than for phosphofructokinase (+cyclic AMP).     

Spectrum of chloroperoxidase compound I

When compound I of chloroperoxidase is formed from the native enzyme the absorption peak in the Soret region diminishes in intensity, and shifts to a maximum absorbance at 367 nm. This unusual Soret spectrum decreases in intensity in a linear fashion as the wavelength increases. The first visible spectrum of chloroperoxidase compound I is reported which has a peak at 689 nm as its most prominent feature.     

Modification by simetryn sulphoxide of a specific thiol group in rat haemoglobin.

Native rat haemoglobins were found to bind simetryn sulphoxide to an extent 40-fold greater than human haemoglobin. This specific behaviour was studied by using only high-pressure ('performance') liquid chromatography for the preparative separation of globin chains and the isolation of peptides resulting from chemical and enzymic degradation. High recoveries (greater than 80%) of peptides throughout the procedures in combination with microsequence techniques, allow a definitive assignment of the residue undergoing modification. The haemoglobin beta-chain cystine-125 residue, with a stoichiometry of one per tetramer of rat haemoglobin, was found to be modified. Stereochemical implications of this finding are discussed. Simetryn sulphoxide would appear to be useful as a specific reagent for the mapping of exposed thiol residues in proteins.     

Cyclosporin A regulates the expression of HLA-DR on human monocytes by two different mechanisms.

Cyclosporin A (CsA), but not its nonimmunosuppressive analog cyclosporin H (CsH), inhibited the expression of HLA-DR in human monocytes. Induction of HLA-DR by interferon (IFN)-gamma in fresh monocytes was also inhibited by CsA and not by CsH. However, when monocytes were pretreated with either CsA or CsH for 16 hr prior to the addition of IFN-gamma, HLA-DR expression was increased, probably because of a cyclosporin-induced increase in the number of IFN-gamma receptors. Down-regulation of the HLA-DR mRNA by CsA was found to be dependent on continuous protein synthesis. IFN-alpha also inhibited the IFN-gamma-induced HLA-DR mRNA expression and showed synergy with CsA at low concentrations but not at high concentrations of the drugs. A common mechanistic element in the pathways of CsA and IFN-alpha is proposed.