Chemical modification of active sites in relation to the catalytic mechanism of F1

Recent studies of chemically modified F₁-ATPases have provided new information that requires a revision of our thinking on their catalytic mechanism. One of the subunits in F₁-ATPase is distinguishable from the other two both structurally and functionally. The catalytic site and regulatory site of the same subunit are probably sufficiently close to each other, and the interaction between the various catalytic and regulatory sites are probably sufficiently strong to raise the uni-site rate of ATP hydrolysis by several orders of magnitude to that of promoted (multi-site) ATP hydrolysis. Although all three subunits in F₁ possess weak uni-site ATPase activity, only one of them () catalyzes promoted ATP hydrolysis. But all three subunits catalyze ATP synthesis driven by the proton flux. Internal rotation of the ₃₃ or ₃ moiety relative to the remainder of the F₀F₁ complex did not occur during oxidative phosphorylation by reconstituted submitochondrial particles.     

Protection of photosynthetic O2 evolution against heat inactivation: the role of chloride,pH and coupling status

Heat inactivation of photosynthetic O₂ evolution was studied in isolated thylakoids from spinach (Spinacia oleracea) and mangrove (Avicennia marina) leaves. Different temperatures, salt, pH and uncoupler effects were investigated. From these results and others in the literature it was concluced that chloride loss from the membrane and, more specifically, the oxygen-evolving complex of photosystem II, may be the cause of inhibition of oxygen evolution during heat inactivation.Abbreviations Hepes 4-(2-hydroxyethyl)-1-piperazinethanesulfonic acid - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol - Tricine N-2-hydroxy-1, 1-bis (hydroxymethyl) ethyl glycine - EDTA ethylenediaminetetraacetic acid - FeCN K-ferricyanide     

Induction of secondary dormancy in sunflower seeds by high temperature. Possible involvement of ethylene biosynthesis

High temperature (45°C) inhibits seed germinition and seedling sunflower ( Helianthus annuus L. cv. Mirasol). Treatment of imbibed seeds at 45°C for more than 48 h induces a secondary dormancy, which is associated with progressive decrease of germination ability at optimal temperature (25°C) as well as with abnormal seedling growth. Ethylene (55μl l⁻¹) and 2-chloroethylphosphonic acid (ethephon) (2.5 m M ) improve germination of thermodormant seeds at 25°C. but the abnormal growth of the seedlings remains. O₂-enriched atmosphere and dry storage improve germination and normal seedling growth. The induction of thermodormancy in sunflower seeds seems associated with loss of their ability to convert 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene. Possible effects of high temperature on membranes and ethylene forming enzyme (EFE) are discussed.     

A rapid non-destructive assay to quantify soybean nodule gas permeability

The low gas permeability of a diffusion barrier in the cortex of soybean nodules plays a significant role in the protection of nitrogenase from oxygen inactivation. It may also set an upper limit on nodule respiration and nitrogen fixation rates. Two methods which have been used to quantify the gas permeability of leguminous nodules are reviewed and found to be unreliable. A new assay technique for determining both the nodule activity and gas permeability is developed and tested. This ‘lag-phase’ assay is based on the time nodules require to reach steady-state ethylene production after being exposed to acetylene. The technique is rapid, insensitive to errors in biochemical parameters associated with nitrogenase, and is non-destructive. The method was tested with intact aeroponically grown soybean plants for which the mean nodule gas permeability was found to be 13.3×10⁻³ mms⁻¹. This corresponds to a layer of cells approximately 35 um thick and is consistent with previously reported values.     

Soybean nodule gas permeability,nitrogen fixation and dirunal cycles in soil temperature

While diurnal cycles in nitrogen fixation rates are sometimes assumed to result from diurnal variation in photosynthetically active radiation, contradicting evidence exists that indicate soil temperature is the primary environmental influence. These studies assessed the significance of temperature on soybean nitrogen fixation under field conditions. Two groups of intact field-grown soybean plants, one at ambient and the other exposed to a 10°C diurnal variation in soil temperature, were nondestructively assayed for acetylene reduction rates. Activity was closely associated with soil temperature (R²=0.85), even when temperature was 12 h out of phase with ambient. Data were also obtained to determine if the effects of rhizosphere temperature on nitrogen fixation are mediated through an effect on the nodule oxygen permeability. Nodule oxygen permeability of intact, aeroponically grown soybean was closely correlated with the diurnal changes in temperature (R²=0.90).     

Plant wound compounds from oviposition scars used in host discrimination by a stem-galling sawfly

The stem-galling sawfly Euura lasiolepisuses one or more plant wound compounds resulting from oviposition scars as cues in host discrimination (avoiding sites occupied by conspecifics). Four experiments were conducted to test hypotheses about how Euurapartitions resources. Experiment 1 demonstrated that Euuraavoids ovipositing on nodes with scars from previous ovipositions. Experiment 2 showed no evidence that the sawfly uses oviposition-deterring pheromones and indicated there is a time lag following oviposition before the oviposition scar becomes a deterrent. Experiment 3 showed that sawflies avoid artificially formed scars, demonstrating that a plant cue alone can lead to host discrimination. Experiment 4 showed that visual or tactile cues are not necessary for host discrimination and indicated that a plant wound compound functions as an oviposition deterrent. Both experimental results and field surveys showed that Euuraoviposition scars were more uniformly distributed than expected if sawflies were ignoring previous ovipositions.     

Xenopus spermatozoon: Is there any correlation between motility and oxygen consumption?

The motility status of Xenopus laevis spermatozoa does not affect their respiration rate. Oxygen consumption for 10⁹ spermatozoa is approximately 0.4 μmol/minute. Oxygen consumption is not increased by gramicidin D, an uncoupler, and it is not blocked by KCN or NaN₃. The adenosine triphosphate (ATP) content of spermatozoa that have been activated is definitely less than that in the spermatozoa that remained immotile. Incubation in KCN, NaN₃, and gramicidin decreases the ATP content and impairs motility. The conclusions of the present study are that in Xenopus spermatozoa motility and oxygen consumption are not correlated, and the composition of the respiratory chain of these spermatozoa presents noteworthy peculiarities.     

色氨酸残基在磷酸烯醇式丙酮酸羧化酶催化功能中的作用

在pH7.5条件下,用NBS对PEP羧化酶中色氨酸残基进行共价修饰表明,PEP羧化酶中48个色氨酸残基均能被NBS修饰。用邹承鲁图解法求得,其中4个残基为酶表现催化活性所必需的。 PEP羧化酶的变构效应剂G6P、Gly及Mal分别与酶预保温后,再经NBS修饰,前两种处理中,同样浓度的NBS所用修饰的色氨酸残基数和处理后的残存酶活与对照相比有很大的差异,而用Mal处理的,两者与对照相差无几。     

Oxygen free radicals in volume overload heart failure

It has been suggested that oxygen free radicals (OFR) depress the excitation-contraction coupling in cardiac muscle. It is possible that a decrease in the cardiac contractility in the failing heart may be due to an increased OFR producing activity of polymorphonuclear (PMN) leukocytes. We studied the OFR producing activity (chemiluminescence) of PMN leukocytes from blood in dogs with heart failure due to chronic volume overload. The animals were divided into two groups: I) normal, (n = 10): II) dogs with mitral insufficiency (MI) of 6 to 9 months duration, (n = 10). Hemodynamic studies were done to establish the presence of heart failure. Blood samples were collected to measure PMN leukocyte chemiluminescence. There was a decrease in the cardiac index and index of myocardial contractility (dp/dt/IIP) and an increase in the left ventricular end-diastolic pressure in dogs with MI indicating left ventricular failure. The peak chemiluminescent activity of the PMN leukocytes in blood of dogs with failure was about four folds greater than that in the blood from normal dogs. These results suggest that there may be an increased OFR generation in dogs with volume overload heart failure. The decrease in the myocardial contractility in the failing heart might be due to an increase in the OFR produced by the PMN leukocytes.     

Singlet oxygen: a potential culprit in myocardial injury?

The purpose of this study was to explore the role of singlet oxygen in cardiovascular injury. To accomplish this objective, we investigated the effect of singlet oxygen [generated from photoactivation of rose-bengal] on the calcium transport and Ca²⁺-ATPase activity of cardiac sarcoplasmic reticulum and compared these results with those obtained by superoxide radical, hydrogen peroxide and hydroxyl radical. Isolated cardiac SR exposed to rose bengal (10 nM) irradiated at (560 nm) produced a significant inhibition of Ca ²⁺ uptake; from 2.27 ± 0.05 to 0.62 ± 0.05 µmol Ca⁺/mg.min (mean ± SE) (P < 0.01) and Ca²⁺-ATPase activity from 2.08 ± 0.05 µmol Pi/min. mg to 0.28 ± 0.04 µmol Pi/min. mg (mean ± SE) (P < 0.01). The inhibition of calcium uptake and Ca²⁺-ATPase activity by rose bengal derived activatedoxygen (singlet oxygen) was dependent on the duration of exposure and intensity of light. The singlet oxygen scavengers ascorbic acid and histidine significantly protected SR Ca²⁺-ATPase against rose bengal derived activated oxygen species but superoxide dismutase and catalase did not attenuate the inhibition. SDS-polyacrylamide gel electrophoresis of SR exposed to photoactivated rose bengal up to 14 min, demonstrated complete loss of Ca²⁺-ATPase monomer band which was significantly protected by histidine. Irradiation of rose bengal also caused an 18% loss of total sulfhydryl groups of SR. On the other hand, superoxide (generated from xanthine oxidase action on xanthine) and hydroxyl radical (0.5 mM H₂O₂ + Fe²⁺ -EDTA) as well as H₂O₂ (12 mM) were without any effect on the 97,000 dalton Ca²⁺-ATPase band ofsarcoplasmic reticulum. The results suggest that oxidative damage of cardiac sarcoplasmic reticulum may be mediated by singlet oxygen. This may represent an important mechanism by which the oxidative injury to the myocardium induces both a loss of tension development and arrhythmogenesis.