Coupling between redox and acid-base energy by cytochrome b-564 in Baker's yeast mitochondria

Baker's yeast mitochondrial cytochrome b-564 is characterized by exhibiting both a labile pH-independent high-potential form (E'o, pH 7 = + 190 mV) and a stable pH-dependent (pKa = 6.8) low-potential form (E'o, pH 7 = + 70 mV). The different behavior of these two forms of cytochrome b-564 versus pH seems to be a decisive factor for transduction of redox energy into acid-base energy in oxidative phosphorylation site 2. Deenergizing treatments, such as ADP plus Pi, result in the conversion of all the mitochondrial cytochrome b-564 into its low-potential form, whereas energization with ATP specifically transforms the cytochrome into its high-potential form, the ATP effect being neutralized by the ATPase inhibitor oligomycin and by the uncoupler FCCP. Accordingly, a minimal model for coupling between redox energy and acid-base energy through an electronically energized and protonated ferricytochrome b-564 intermediate is proposed. The energy-transducing properties of mitochondrial cytochrome b-564 seems to be shared by chloroplast cytochrome b-559.     

Inhibition of ceruloplasmin and other copper oxidases by thiomolybdate

The dietary antagonism between copper and molybdate salts prompted a study of the inhibition of copper enzymes by thiomolybdate (TM). TM strongly inhibited the oxidase activity of five copper oxidase with I50% values in the 1-5 microM range. The mechanism of the TM effect on the copper oxidase, ceruloplasmin (Cp) (E.C. 1.16.3.1), was studied in detail. In Vmax vs. E plots, TM gave parallel data suggesting irreversibility but a large number of TM molecules per Cp were required. The inhibition of Cp by TM could not be reversed by dialysis. Isolation of TM-inhibited Cp on Sephadex G-10 did not yield any active Cp molecules. Cu(II) did not restore any inhibited oxidase activity. Gel electrophoresis supported the covalent binding of Cp by TM without any extensive change in protein structure. EPR results confirmed that Cu(II) is reduced to Cu(I) after reaction with TM. However, the Mo(VI) in MoS4(2-) did not change in oxidation number. Analysis of the TM-Cp compound accounted for all six Cu atoms as found in native Cp. The data suggest the covalent binding of sulfide to Cp copper. TM also inhibited the activity of ascorbate oxidase, cytochrome oxidase, superoxide dismutase, and tyrosinase. However, no inhibition of carbonic anhydrase, a zinc enzyme, was observed at 1 mM TM.     

Methionine adenosyltransferase activity in cultured cells and in human tissues

We have investigated methionine adenosyltransferase activity (MAT) in extracts of a variety of normal and malignant human tissues and cultured cell lines. MAT activity assayed from 17 different cultured cell lines varied to a great extent. Ramos (human, Burkitt's lymphoma) and EL4 (mouse, T cell lymphoma) cell showed MAT activity near 300 pmol/mg per min. Daudi (human, Burkitt's lymphoma) and almost all monolayer cells had MAT activity below 100 pmol/mg per min. Human peripheral blood lymphocytes had MAT activity of 36 pmol/mg permin. The MAT activity of the cell lines can be related to doubling time: cell lines with short doubling times have much higher MAT activity than other cell lines. A large variation in MAT activity in different human tissues was observed. In autopsy samples MAT activity was highest in the brain and in the colon. Malignant tissue samples gave much higher MAT activity than normal tissues. Lung cancer (carcinoma squamocellulare pulmonis) had MAT activity of 30.7 pmol/mg per min, while in normal lung it was 2.4 pmol/mg per min.     

Regulation of the binding of 7,12-dimethylbenz[a]-anthracene to DNA of cultured murine epidermal cells at metabolic level: competition of the binding by polycyclic aromatic hydrocarbons and by other precarcinogens.

The binding of labeled carcinogen [3H]DMBA to murine epidermal cells (MEC) DNA in culture has been studied. The influence of unlabeled noncarcinogenic and carcinogenic polycyclic aromatic hydrocarbons (PAH), several PAH metablites, and various directly and indirectly acting non-PAH carcinogens on the binding of [3H]DMBA to MEC DNA has been examined. All the carcinogenic PAH and some of non-carcinogenic PAH effectively inhibit the binding of [3H]DMBA to MEC DNA. The non-PAH chemical carcinogens requiring metabolic activation also reduce the binding of labeled DMBA to MEC DNA; however, a higher concentration of these compounds is required for 50% inhibition of binding than the concentrations of PAH for the same degree of inhibition of binding of [3H]DMBA to MEC DNA. The directly acting carcinogens do not significantly inhibit the binding of [3H]DMBA to DNA. The relationship between structures of PAH and their ability to inhibit the binding of [3H]DMBA to MEC DNA is also discussed. Thus, it appears that the binding of DMBA to cellular DNA is primarily controlled at a level of metabolism and to some extent at the level of binding of reactive metabolites to DNA.     

The subunit structure of mouse satellite chromatin.

Atomic coordinates are presented for the 3740 atoms other than hydrogen in the dimeric molecule of chicken muscle triose phosphate isomerase. They are derived from an electron-density map at 2.5Åresolution, interpreted in terms of the known amino-acid sequence, and they have been adjusted systematically to give stereochemically appropriate bond lengths and angles.     

Crystal structure of the kainate receptor GluR5 ligand-binding core in complex with (S)-glutamate

The X-ray structure of the ligand-binding core of the kainate receptor GluR5 (GluR5-S1S2) in complex with (S)-glutamate was determined to 1.95 A resolution. The overall GluR5-S1S2 structure comprises two domains and is similar to the related AMPA receptor GluR2-S1S2J. (S)-glutamate binds as in GluR2-S1S2J. Distinct features are observed for Ser741, which stabilizes a highly coordinated network of water molecules and forms an interdomain bridge. The GluR5 complex exhibits a high degree of domain closure (26 degrees) relative to apo GluR2-S1S2J. In addition, GluR5-S1S2 forms a novel dimer interface with a different arrangement of the two protomers compared to GluR2-S1S2J.     

Mixed-ligand complex formation of mercury (II) ethylenediaminetetraacetate with cysteine and methionine in aqueous solution

The mixed-ligand complex formation in the systems Hg²⁺-Edta⁴⁻-L (L = Cys²⁻, Met⁻) has been studied by means of calorimetry, pH-potentiometry and NMR spectroscopy in aqueous solution at 298.15 K and the ionic strength of I = 0.5 (KNO₃). The thermodynamic parameters of formation of the HgEdtaL, HgEdtaHL and (HgEdta)₂L complexes have been determined. The most probable coordination mode for the complexone and the amino acid in the mixed-ligand complexes is discussed.     

Possible effects of the detachment of stromal lamellae from granal stacks on salt-induced changes in spillover. A study by sonication of chloroplasts

Salt-induced chlorophyll fluorescence and spillover changes in control and briefly sonicated chloroplasts have been studied under conditions where Photosystem II traps are closed. In a low-salt medium containing 10 mM KCl, control envelope-free chloroplasts exhibited good spillover, as measured by low chlorophyll fluorescence yield at room temperature, a high ratio of the fluorescence peaks $\text{F}{}_{735}\text{F}{}_{685}$ at 77 K, and increased Photosystem I activity in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea and Photosystem II light. In contrast, when stacked chloroplasts were briefly sonicated and subsequently diluted into a low-salt medium, a high fluorescence yield at room temperature and a low ratio of $\text{F}{}_{735}\text{F}{}_{685}$ at 77 K persisted. When unstacked chloroplasts were sonicated and then diluted into a high-salt medium, the room temperature fluorescence yield remained low. The results are interpreted in terms of a model relating the changes in chlorophyll fluoresecence with the lateral diffusion of Photosystem I and Photosystem II chlorophyll-protein complexes in the plane of the thylakoid membrane creating randomized or segregated domains, depending on the degree of electrostatic screening of surface charges (Barber, J. (1980) FEBS Lett. 188, 1–10). It is argued that brief sonication of stacked chloroplasts separates stromal membranes from granal stacks, thus limiting the inter-mixing of the photosystems via lateral diffusion even when the ionic composition of the medium is varied. Consequently energy transfer from Photosystem II to Photosystem I is relatively poor and chlorophyll fluorescence from Photosystem II is enhanced. The loss of the salt effect on sonicated unstacked membranes can also be accommodated by the model. In this case it seems that the generation of small membrane fragments does not allow the normal salt-induced phase separation of the pigment-protein complexes to occur.     

Strictinin as an efficient antioxidant in lipid peroxidation

The antioxidant effect of strictinin (SOH), which was extracted from green tea leaves, against the peroxidation of linoleic acid in sodium dodecyl sulfate (SDS) and cetyl trimethylammonium (CTAB) micelles, against the peroxidation of low-density lipoprotein (LDL) and against oxidative hemolysis of human red blood cells (RBCs), has been studied. The peroxidation of linoleic acid and LDL, and oxidative hemolysis of RBCs were initiated thermally by a water-soluble azo initiator 2,2'-azobis(2-amidinopropane hydrochloride) (AAPH), and the reaction kinetics in micelles and LDL were monitored by uptake of oxygen. The synergistic antioxidant effect of SOH with alpha-tocopherol (Vitamin E) was also studied by following the decay kinetics of alpha-tocopherol. Kinetic analysis of the antioxidation process demonstrates that SOH, used either alone or in combination with alpha-tocopherol, is an effective antioxidant against lipid peroxidation, but its effects significantly depend on the reaction medium.