Site Of ATP Synthesis Research Articles

AMP-Forming Acetyl Coenzyme A Synthetase in the Outermost Membrane of the Hyperthermophilic Crenarchaeon Ignicoccus hospitalis

Ignicoccus hospitalis, a hyperthermophilic, chemolithoautotrophic crenarchaeon was found to possess a new CO(2) fixation pathway, the dicarboxylate/4-hydroxybutyrate cycle. The primary acceptor molecule for this pathway is acetyl coenzyme A (acetyl-CoA), which is regenerated in the cycle via the characteristic intermediate 4-hydroxybutyrate. In the presence of acetate, acetyl-CoA can alternatively be formed in a one-step mechanism via an AMP-forming acetyl-CoA synthetase (ACS). This enzyme was identified after membrane preparation by two-dimensional native PAGE/SDS-PAGE, followed by matrix-assisted laser desorption ionization-time of flight tandem mass spectrometry and N-terminal sequencing. The ACS of I. hospitalis exhibits a molecular mass of ∼690 kDa with a monomeric molecular mass of 77 kDa. Activity tests on isolated membranes and bioinformatic analyses indicated that the ACS is a constitutive membrane-associated (but not an integral) protein complex. Unexpectedly, immunolabeling on cells of I. hospitalis and other described Ignicoccus species revealed that the ACS is localized at the outermost membrane. This perfectly coincides with recent results that the ATP synthase and the H(2):sulfur oxidoreductase complexes are also located in the outermost membrane of I. hospitalis. These results imply that the intermembrane compartment of I. hospitalis is not only the site of ATP synthesis but may also be involved in the primary steps of CO(2) fixation.

The Coronary Circulation in Arterial Hypertension

Arterial hypertension is a major coronary risk factor due to an increase prevalence of coronary atherosclerosis. On the other hand, many arguments are suggestive of symptomatic or silent episodes of myocardial ischemia in hypertensive patients without coronary artery stenosis. Several pathophysiological processes that are implicated in coronary vascular changes are detrimental for the oxygen demand/supply equilibrium and adaptation of coronary circulation and coronary blood flow to an increase in myocardial oxygen demand. First, apart from coronary atherosclerosis, epicardial vessels are altered by vessel wall remodelling and structure changes. Second, structural alterations of the coronary arterioles due to medial hypertrophy and perivascular fibrosis limit the functional area of the vessels which are also compressed by a higher intramyocardial pressure, resulting in a reduction of coronary flow reserve. Third, the increase in intercapillary distance due to myocardial hypertrophy increases the distance of oxygen diffusion to cardiac myocyte mitochondria, the site of ATP synthesis. Fourth, the endothelium-dependent nitric-oxide dilation of coronary vessels is depressed, thus impairing the adaptation of coronary circulation to changes in myocardial oxygen demand. All these changes do not depend on myocardial hypertrophy, although myocardial hypertrophy aggravates the disorders. Antihypertensive therapy may have beneficial effects by reducing myocardial mass and arterial pressure, by restoring a normal vascular structure and architecture, and by improving coronary vasomotion. Keywords: Arterial hypertension, Coronary circulation, Vascular structure, Vascular function, Antihypertensive therapy

Focally Elevated Creatine Detected in Amyloid Precursor Protein (APP) Transgenic Mice and Alzheimer Disease Brain Tissue

The creatine/phosphocreatine system, regulated by creatine kinase, plays an important role in maintaining energy balance in the brain. Energy metabolism and the function of creatine kinase are known to be affected in Alzheimer diseased brain and in cells exposed to the beta-amyloid peptide. We used infrared microspectroscopy to examine hippocampal, cortical, and caudal tissue from 21-89-week-old transgenic mice expressing doubly mutant (K670N/M671L and V717F) amyloid precursor protein and displaying robust pathology from an early age. Microcrystalline deposits of creatine, suggestive of perturbed energetic status, were detected by infrared microspectroscopy in all animals with advanced plaque pathology. Relatively large creatine deposits were also found in hippocampal sections from post-mortem Alzheimer diseased human brain, compared with hippocampus from non-demented brain. We therefore speculate that this molecule is a marker of the disease process.

The site of oxygen limitation in soybean nodules

In legume nodules the [O2] in the infected cells limits respiration and nitrogenase activity, becoming more severe if nodules are exposed to subambient O2 levels. To identify the site of O2 limitation, adenylate pools were measured in soybean (Glycine max) nodules that were frozen in liquid N2 before being ground, lyophilized, sonicated, and separated on density gradients of nonaqueous solvents (heptane/tetrachloroethylene) to yield fractions enriched in bacteroid or plant components. In nodules maintained in air, the adenylate energy charge (AEC = [ATP + 0.5 ADP]/[ATP + ADP + AMP]) was lower in the plant compartment (0.65 +/- 0.04) than in the bacteroids (0.76 +/- 0.095), but did not change when the nodulated root system was exposed to 10% O2. In contrast, 10% O2 decreased the bacteroid AEC to 0.56 +/- 0.06, leading to the conclusion that they are the primary site of O2 limitation in nodules. To account for the low but unchanged AEC in the plant compartment and for the evidence that mitochondria are localized in O2-enriched microenvironments adjacent to intercellular spaces, we propose that steep adenylate gradients may exist between the site of ATP synthesis (and ADP use) in the mitochondria and the extra-mitochondrial sites of ATP use (and ADP production) throughout the large, infected cells.

Functional and Evolutionary Implications of the Distribution of Phosphagens in Primitive-Type Spermatozoa.

External fertilization is considered to be the primitive condition in metazoans. The spermatozoa of such organisms typically display a common primitive-type morphology that is present in a range of phyla. These spermatozoa are extremely polarized cells in that the site of ATP synthesis (mitochondria in midpiece) is located at large diffusion distances from the ATP sink (dynein ATPases in the flagellum). Spermatozoa of polychaetes, sipunculids, echiuroids, echinoderms, and tunicates contain the phosphagen creatine phosphate or express the corresponding phosphagen kinase creatine kinase (or both), even when other phosphagens/phosphagen kinases are present in somatic tissues and eggs. The selective expression of the creatine kinase system in these spermatozoa may be related to potential advantages in the cellular transport of energy. To evaluate this possibility, we compared the efficacy of the major phosphagen systems for cellular transport of energy. We used a facilitated diffusion model for spatial ATP buffering, taking into account relative differences in diffusivity and thermodynamic poise. At low ratios of [total phosphagen pool]/ [total adenine nucleotide pool] (CG+P/CAd ratio), creatine phosphate carried a higher fraction of total high-energy phosphate (J) than the other phosphagens. However, J values for all phosphagens were greater than 0.9, and these differences disappeared as the CG+P/CAd ratio was increased. Thus, the functional benefit of using CP, rather than other phosphagens, in energy transport is quite limited. The creatine kinase system became associated with primitive-type spermatozoa early in metazoan evolution. This association is not necessarily related to inherent advantages of this phosphagen system for buffering of ATP, but may be linked to historical events in the evolution of the cell phenotype.

Acetic acid from H2 and CO2

Cell extracts of a nonsporeforming strictly anaerobic bacterium, Acetobacterium woodii produced acetate in N-tris(Hydroxymethyl)methyl-2-aminoethane sulfonic acid or phosphate buffers from hydrogen and carbon dioxide. The formation of acetate was not dependent on the presence of ATP in the reaction mixture; ADP also did not influence the acetate production. Since acetic acid is the main fermentation product during growth of A. woodii with H2 and CO2, ATP must be synthesized in the course of acetate formation. The possible sites of ATP synthesis are discussed.

Evidence for chemiosmotic coupling of electron transport to ATP synthesis in spinach chloroplasts

In spinach chloroplasts it has been shown that (1) the size of the proton gradient under phosphorylating conditions is smaller than under non-phosphorylating conditions; (2) ADP, ATP or Dio-9, added under non-phosphorylating conditions, decrease the rate of electron transport but increase the size of the proton gradient; (3) ADP, ATP or Dio-9 inhibit not only electron transport but also the rate of decay of the proton gradient; (4) the H + e − ratio under non-phosphorylating conditions is 1.0. It is not affected by ADP, ATP or Dio-9. These results show that protons pass out of the thylakoids at the site of ATP synthesis and that this leakage is inhibited by ADP, ATP or Dio-9, compounds that interact with the site of ATP synthesis. As these compounds do not alter the H + e − ratio the formation of the proton gradient must be an intermediate between electron transport and ATP synthesis. These data are in support of the chemiosmotic theory of coupling of electron transport to ATP synthesis.

Inhibitors and uncouplers of photophosphorylation

Abstract 1. 1. A survey of the effect of inhibitors and uncouplers of oxidative phosphorylation on chloroplast photophosphorylation led to the discovery of new inhibitors and uncouplers of this process. 2. 2. n-Butyl-3,5-diiodo-4-hydroxybenzoate was found to inhibit both electron transport and the coupled ATP formation. However, phenazine methosulfate-catalyzed photophosphorylation was much less inhibited than all the other photoreactions tested. 3. 3. By analogy with the inhibition caused by 3-(3,4-dichlorophenyl)-1,1-dimethylurea, it is suggested that n-butyl-3,5-diiodo-4-hydroxybenzoate effects steps related to the light reaction of System II. 4. 4. Several compounds were found to act as uncouplers of photophosphorylation: atebrin, chlorpromazine, gramicidin S, octyl guanidine and thiophosphate. 5. 5. Preferential uncoupling of the ferricyanide-dependent phosphorylation was found with the following uncouplers: carbonyl cyanide p-trifluoromethoxyphenylhydrazone, octyl guanidine and gramicidin S. This differential uncoupling suggests the existence of two different sites of ATP synthesis. 6. 6. Some inhibitors and uncouplers became more effective and others less effective when the light intensity was lowered. These opposite effects indicate that some of these agents act at earlier steps than others. 7. 7. No significant inhibition was given by rotenone, aurovertin, chlortetracycline, oxytetracycline, arsenite and Tl3+ ions. Oligomycin and valinomycin acted as weak uncouplers at relatively high concentrations.