DNA-dependent RNA polymerase from Spirochaeta aurantia

Abstract A DNA-dependent RNA polymerase was isolated from Spirochaeta aurantia . The M _r values of the holoenzyme subunits are 164000, 142000, 84000, and 44500. The RNA polymerase activity was sensitive to heparin, streptolydigin, and actinomycin D, while rifampicin and streptovaricin did not inhibit activity.     

Effect of b-c1-site inhibitors on the midpoint potentials of mitochondrial cytochromes b

Anaerobic potentiometric titrations of b cytochromes have been carried out in beef heart submitochondrial particles in the presence of several specific inhibitors of electron transfer through the b-c1-site of the respiratory chain. Whereas antimycin shows no significant effect on the titration curve of cytochrome b-562, NoHOQnO is found to shift the Em of b-562 by 20-30 mV to the positive. Funiculosin raises the Em of b-562 by greater than 100 mV and also appears to bring about a minor shift of b-566 midpoint potential. In the presence of myxothiazol, both b cytochromes titrate with Em values 15-30 mV more positive than in the control.     

Interaction of nucleic acids with the DNA-dependent RNA polymerases of Drosophila

The interaction between the three Drosophila DNA-dependent RNA polymerases (EC 2.7.7.6) and the DNA template or the RNA product was investigated by photochemical cross-linking and binding studies, using RNA polymerase subunits immobilized on nitro-cellulose filters. It can be shown that the two largest subunits are responsible for the binding of the enzymes to both template and newly-synthesized RNA.     

Streptozotocin diabetes results in increased responsiveness of adipocyte lipolysis to glucagon

Adipocytes from streptozotocin-diabetic rats are approximately 50-times more sensitive to the lipolytic action of glucagon. This change is only perceived in the presence of a small quantity of adenosine deaminase which itself has little effect on basal lipolysis. Insulin treatment restores glucagon sensitivity to normal.     

Changes in composition and function of thylakoid membranes as a result of photosynthetic adaptation of chloroplasts from pea plants grown under different light conditions

The hypothesis that chloroplasts having different light-saturated rates of photosynthesis will have different proportions of the intrinsic thylakoid complexes engaged in light-harvesting and electron transport (Anderson, J.M. (1982) Mol. Cell. Biochem. 46, 161–172) has been tested. Peas were grown in light regimes which varied in light intensity, quality and time of irradiance, and ranged from sunlight through red to blue-enriched light of very low radiation. The electron-transport capacity at saturating light of Photosystem I and Photosystem II of chloroplasts isolated from light-adapted peas was 2-fold and 5–6-fold lower, respectively, in the lowest radiation compared to sunlight. There was a marked increase in the amount of total chlorophyll associated with the main chlorophyll $\text{a}\text{b-}\text{proteins}$ (LHCP¹, LHCP² and LHCP³) and a 2-fold decrease in the core reaction centre complex of Photosystem II (CP a) as the radiation decreased; the $\text{LHCP}{}^{\mathrm{1–3}}\text{CP}$ a ratio changed from 3.5 to 9.0. The amount of chlorophyll associated with Photosystem I varied from 34% in sunlight to 27% in the lowest radiation, but the antenna size of Photosystem I was not markedly different; there was a 2-fold decrease in the amount of cytochrome f on a chlorophyll basis, which partly accounted for the decreased electron-transport capacity of Photosystem I. Since the increases or decreases in the levels of each of the components correlated with decreasing radiation, it is clear that the light-adaptation of both light-harvesting and electron-transport components is indeed closely co-ordinated.     

Effects of N,N′-dicyclohexylcarbodiimide on isolated and reconstituted cytochrome b-c1 complex from bovine heart mitochondria

N,N′-Dicyclohexylcarbodiimide (DCCD) inhibits the activity of ubiquinol-cytochrome c reductase in the isolated and reconstitued mitochondrial cytochrome b-c₁ complex. DCCD inhibits equally electron flow and proton translocation (i.e., the $\text{H}{}^{\mathrm{+}}\text{e}{}^{\mathrm{?}}$ ratio is not affected) catalysed by the enzyme reconstituted into phospholipid vesicles. The inhibitory effects are accompanied by structural alterations in the polypeptide pattern of both isolated and reconstituted enzyme. Cross-linking was observed between subunits V (iron-sulfur protein) and VII, indicating that these polypeptides are in close proximity. A clear correlation was found between the kinetics of inhibition of enzymic activity and the cross-linking, suggesting that the two phenomena may be coupled. Binding of [¹⁴C]DCCD was also observed, to all subunits with the isolated enzyme and preferentially to cytochrome b with the reconstituted vesicles; in both cases, however, it was not correlated kinetically with the inhibition of the enzymic activity.     

Impedance of the electrogenic Cl− pump inAcetabularia: Electrical frequency entrainements,voltage-sensitivity,and reaction kinetic interpretation

Summary Reaction kinetic analysis of the electrical properties of the electrogenic Cl^– pump inAcetabularia has been extended from steady-state to nonsteady-state conditions: electrical frequency responses of theAcetabularia membrane have been measured over the range from 1 Hz to 10 kHz at transmembrane potential differences across the plasmalemma (V _m ) between –70 and –240 mV using voltage-clamp techniques. The results are well described by an electrical equivalent circuit with three parallel limbs: a conventional membrane capacitancec _m , a steadystate conductanceg _o (predominantly of the pump pathway plus a minor passive ion conductance) and a conductanceg _s in series with a capacitancec _p which are peculiar to the temporal behavior of the pump. The absolute values and voltage sensitivities of these four elements have been determined:c _m of about 8 mF m^–2 turned out to be voltage insensitive; it is considered to be normal.g _o is voltage sensitive and displays a peak of about 80 S m^–2 around –180 mV. Voltage sensitivity ofg _s could not be documented due to large scatter ofg _s (around 80 S m^–2).c _p behaved voltage sensitive with a notch of about 20 mF m^–2 around –180 mV, a peak of about 40 mF m^–2 at –120 mV and vanishing at –70 mV. When these data are compared with the predictions of nonsteady-state electrical properties of charge transport systems (U.-P. Hansen, J. Tittor, D. Gradmann, 1983,J. Membrane Biol. in press), model A (redistribution of states within the reaction cycle) consistently provides magnitude and voltage sensitivity of the elementsg _o ,g _s andc _p of the equivalent circuit, when known kinetic parameters of the pump are used for the calculations. This analysis results in a density of pump elements in theAcetabularia plasmalemma of about 50 nmol m^–2. The dominating rate constants for the redistribution of the individual states of the pump in the electric field turn out to be in the range of 500 sec^–1, under normal conditions.     

Purification and properties of a low molecular weight DNA polymerase from Neurospora crassa

A third DNA polymerase ‘C’ with low molecular weight was isolated and purified 3700-fold from ground hyphae of Neurospora crassa WT 74 A, which shows similarities to β- and γ-polymerases from higher eukaryotes: preference for poly(rA)(dT) as a template/primer, inhibition by p-chloromercuribenzoate, resistance against N-ethylmaleimide up to 10 mmol/l, and molecular weight of about 40 000. This polymerase elutes as a distinct peak from DEAE-cellulose at 0.60 mol/l KCl and has an optimum for K⁺ at 2–20 mmol/l, for Mn²⁺ at 0.8 mmol/l, for Mg²⁺ at 4.0 mmol/l, the pH optimum is 8.0. Its K_m is 1.5 μmol/l using dTTP as substrate. The enzyme activity described here is free of endonuclease but contains detectable amounts of exonuclease.