Calcium and copper transport ATPases: analogies and diversities in transduction and signaling mechanisms

The calcium transport ATPase and the copper transport ATPase are members of the P-ATPase family and retain an analogous catalytic mechanism for ATP utilization, including intermediate phosphoryl transfer to a conserved aspartyl residue, vectorial displacement of bound cation, and final hydrolytic cleavage of Pi. Both ATPases undergo protein conformational changes concomitant with catalytic events. Yet, the two ATPases are prototypes of different features with regard to transduction and signaling mechanisms. The calcium ATPase resides stably on membranes delimiting cellular compartments, acquires free Ca2+ with high affinity on one side of the membrane, and releases the bound Ca2+ on the other side of the membrane to yield a high free Ca2+ gradient. These features are a basic requirement for cellular Ca2+ signaling mechanisms. On the other hand, the copper ATPase acquires copper through exchange with donor proteins, and undergoes intracellular trafficking to deliver copper to acceptor proteins. In addition to the cation transport site and the conserved aspartate undergoing catalytic phosphorylation, the copper ATPase has copper binding regulatory sites on a unique N-terminal protein extension, and has also serine residues undergoing kinase assisted phosphorylation. These additional features are involved in the mechanism of copper ATPase intracellular trafficking which is required to deliver copper to plasma membranes for extrusion, and to the trans-Golgi network for incorporation into metalloproteins. Isoform specific glyocosylation contributes to stabilization of ATP7A copper ATPase in plasma membranes

Atomic-Level Characterization of the Activation Mechanism of SERCA by Calcium

We have performed molecular dynamics (MD) simulations to elucidate, in atomic detail, the mechanism by which the sarcoplasmic reticulum Ca2+-ATPase (SERCA) is activated by Ca2+. Crystal structures suggest that activation of SERCA occurs when the cytoplasmic head-piece, in an open (E1) conformation stabilized by Ca2+, undergoes a large-scale open-to-closed (E1 to E2) transition that is induced by ATP binding. However, spectroscopic measurements in solution suggest that these structural states (E1 and E2) are not tightly coupled to biochemical states (defined by bound ligands); the closed E2 state predominates even in the absence of ATP, in both the presence and absence of Ca2+. How is this loose coupling consistent with the high efficiency of energy transduction in the Ca2+-ATPase? To provide insight into this question, we performed long (500 ns) all-atom MD simulations starting from the open crystal structure, including a lipid bilayer and water. In both the presence and absence of Ca2+, we observed a large-scale open-to-closed conformational transition within 400 ns, supporting the weak coupling between structural and biochemical states. However, upon closer inspection, it is clear that Ca2+ is necessary and sufficient for SERCA to reach the precise geometrical arrangement necessary for activation of ATP hydrolysis. Contrary to suggestions from crystal structures, but in agreement with solution spectroscopy, the presence of ATP is not required for this activating transition. Principal component analysis showed that Ca2+ reshapes the free energy landscape of SERCA to create a path between the open conformation and the activated closed conformation. Thus the malleability of the free energy landscape is essential for SERCA efficiency, ensuring that ATP hydrolysis is tightly coupled to Ca2+ transport. These results demonstrate the importance of real-time dynamics in the formation of catalytically competent conformations of SERCA, with broad implications for understanding enzymatic catalysis in atomic detail

Augmented Cardiac Hypertrophy in Response to Pressure Overload in Mice Lacking ELTD1

BACKGROUND: Epidermal growth factor (EGF), latrophilin and seven transmembrane domain-containing protein 1 (ELTD1) is developmentally upregulated in the heart. Little is known about the relationship between ELTD1 and cardiac diseases. Therefore, we aimed to clarify the role of ELTD1 in pressure overload-induced cardiac hypertrophy. METHODS AND RESULTS: C57BL/6J wild-type (WT) mice and ELTD1-knockout (KO) mice were subjected to left ventricular pressure overload by descending aortic banding (AB). KO mice exhibited more unfavorable cardiac remodeling than WT mice 28 days post AB; this remodeling was characterized by aggravated cardiomyocyte hypertrophy, thickening of the ventricular walls, dilated chambers, increased fibrosis, and blunted systolic and diastolic cardiac function. Analysis of signaling pathways revealed enhanced extracellular signal-regulated kinase (ERK) and the c-Jun amino-terminal kinase (JNK) phosphorylation in response to ELTD1 deletion. CONCLUSIONS: ELTD1 deficiency exacerbates cardiac hypertrophy and cardiac function induced by AB-induced pressure overload by promoting both cardiomyocyte hypertrophy and cardiac fibrosis. These effects are suggested to originate from the activation of the ERK and JNK pathways, suggesting that ELTD1 is a potential target for therapies that prevent the development of cardiac disease

Catalysis over zinc-incorporated berlinite (ZnAlPO4) of the methoxycarbonylation of 1,6-hexanediamine with dimethyl carbonate to form dimethylhexane-1,6-dicarbamate

<p>Abstract</p> <p>Background</p> <p>The alkoxycarbonylation of diamines with dialkyl carbonates presents promising route for the synthesis of dicarbamates, one that is potentially 'greener' owing to the lack of a reliance on phosgene. While a few homogeneous catalysts have been reported, no heterogeneous catalyst could be found in the literature for use in the synthesis of dicarbamates from diamines and dialkyl carbonates. Because heterogeneous catalysts are more manageable than homogeneous catalysts as regards separation and recycling, in our study, we hydrothermally synthesized and used pure berlinite (AlPO₄) and zinc-incorporated berlinite (ZnAlPO₄) as heterogeneous catalysts in the production of dimethylhexane-1,6-dicarbamate from 1,6-hexanediamine (HDA) and dimethyl carbonate (DMC). The catalysts were characterized by means of XRD, FT-IR and XPS. Various influencing factors, such as the HDA/DMC molar ratio, reaction temperature, reaction time, and ZnAlPO₄/HDA ratio, were investigated systematically.</p> <p>Results</p> <p>The XRD characterization identified a berlinite structure associated with both the AlPO₄and ZnAlPO₄catalysts. The FT-IR result confirmed the incorporation of zinc into the berlinite framework for ZnAlPO₄. The XPS measurement revealed that the zinc ions in the ZnAlPO₄structure possessed a higher binding energy than those in ZnO, and as a result, a greater electron-attracting ability. It was found that ZnAlPO₄catalyzed the formation of dimethylhexane-1,6-dicarbamate from the methoxycarbonylation of HDA with DMC, while no activity was detected on using AlPO₄. Under optimum reaction conditions (i.e. a DMC/HDA molar ratio of 8:1, reaction temperature of 349 K, reaction time of 8 h, and ZnAlPO₄/HDA ratio of 5 (mg/mmol)), a yield of up to 92.5% of dimethylhexane-1,6-dicarbamate (with almost 100% conversion of HDA) was obtained. Based on these results, a possible mechanism for the methoxycarbonylation over ZnAlPO₄was also proposed.</p> <p>Conclusion</p> <p>As a heterogeneous catalyst ZnAlPO₄berlinite is highly active and selective for the methoxycarbonylation of HDA with DMC. We propose that dimethylhexane-1,6-dicarbamate is formed <it>via </it>a catalytic cycle, which involves activation of the DMC by a key active intermediate species, formed from the coordination of the carbonyl oxygen with Zn(II), as well as a reaction intermediate formed from the nucleophilic attack of the amino group on the carbonyl carbon.</p

Diabetes Alters Intracellular Calcium Transients in Cardiac Endothelial Cells

Diabetic cardiomyopathy (DCM) is a diabetic complication, which results in myocardial dysfunction independent of other etiological factors. Abnormal intracellular calcium ([Ca2+]i) homeostasis has been implicated in DCM and may precede clinical manifestation. Studies in cardiomyocytes have shown that diabetes results in impaired [Ca2+]i homeostasis due to altered sarcoplasmic reticulum Ca2+ ATPase (SERCA) and sodium-calcium exchanger (NCX) activity. Importantly, altered calcium homeostasis may also be involved in diabetes-associated endothelial dysfunction, including impaired endothelium-dependent relaxation and a diminished capacity to generate nitric oxide (NO), elevated cell adhesion molecules, and decreased angiogenic growth factors. However, the effect of diabetes on Ca2+ regulatory mechanisms in cardiac endothelial cells (CECs) remains unknown. The objective of this study was to determine the effect of diabetes on [Ca2+]i homeostasis in CECs in the rat model (streptozotocin-induced) of DCM. DCM-associated cardiac fibrosis was confirmed using picrosirius red staining of the myocardium. CECs isolated from the myocardium of diabetic and wild-type rats were loaded with Fura-2, and UTP-evoked [Ca2+]i transients were compared under various combinations of SERCA, sarcoplasmic reticulum Ca2+ ATPase (PMCA) and NCX inhibitors. Diabetes resulted in significant alterations in SERCA and NCX activities in CECs during [Ca2+]i sequestration and efflux, respectively, while no difference in PMCA activity between diabetic and wild-type cells was observed. These results improve our understanding of how diabetes affects calcium regulation in CECs, and may contribute to the development of new therapies for DCM treatment

Mice Null for Calsequestrin 1 Exhibit Deficits in Functional Performance and Sarcoplasmic Reticulum Calcium Handling

In skeletal muscle, the release of calcium (Ca2+) by ryanodine sensitive sarcoplasmic reticulum (SR) Ca2+ release channels (i.e., ryanodine receptors; RyR1s) is the primary determinant of contractile filament activation. Much attention has been focused on calsequestrin (CASQ1) and its role in SR Ca2+ buffering as well as its potential for modulating RyR1, the L-type Ca2+ channel (dihydropyridine receptor, DHPR) and other sarcolemmal channels through sensing luminal [Ca2+]. The genetic ablation of CASQ1 expression results in significant alterations in SR Ca2+ content and SR Ca2+ release especially during prolonged activation. While these findings predict a significant loss-of-function phenotype in vivo, little information on functional status of CASQ1 null mice is available. We examined fast muscle in vivo and in vitro and identified significant deficits in functional performance that indicate an inability to sustain contractile activation. In single CASQ1 null skeletal myofibers we demonstrate a decrease in voltage dependent RyR Ca2+ release with single action potentials and a collapse of the Ca2+ release with repetitive trains. Under voltage clamp, SR Ca2+ release flux and total SR Ca2+ release are significantly reduced in CASQ1 null myofibers. The decrease in peak Ca2+ release flux appears to be solely due to elimination of the slowly decaying component of SR Ca2+ release, whereas the rapidly decaying component of SR Ca2+ release is not altered in either amplitude or time course in CASQ1 null fibers. Finally, intra-SR [Ca2+] during ligand and voltage activation of RyR1 revealed a significant decrease in the SR[Ca2+]free in intact CASQ1 null fibers and a increase in the release and uptake kinetics consistent with a depletion of intra-SR Ca2+ buffering capacity. Taken together we have revealed that the genetic ablation of CASQ1 expression results in significant functional deficits consistent with a decrease in the slowly decaying component of SR Ca2+ release

The role of power in financial statement fraud schemes

In this paper, we investigate a large-scale financial statement fraud to better understand the process by which individuals are recruited to participate in financial statement fraud schemes. The case reveals that perpetrators often use power to recruit others to participate in fraudulent acts. To illustrate how power is used, we propose a model, based upon the classical French and Raven taxonomy of power, that explains how one individual influences another individual to participate in financial statement fraud. We also provide propositions for future research

Acyloxylation of Cyclic Enones: Synthesis of Densely Oxygenated Guaianolides

The α′-acyloxylation of cyclic enones with linear carboxylic acids is described. The reaction is promoted by KMnO4 in the presence of a carboxylic acid and its corresponding carboxylic anhydride. The optimization of the reaction has been carried out using the statistical methodology known as design of experiments. The optimized reaction conditions have been evaluated in terms of substrate scope and compatibility with different functional groups. The methodology has been applied to the synthesis of densely oxygenated guaianes and guaianolides

Flavonoids in prevention of diseases with respect to modulation of Ca-pump function

Flavonoids, natural phenolic compounds, are known as agents with strong antioxidant properties. In many diseases associated with oxidative/nitrosative stress and aging they provide multiple biological health benefits. Ca2+-ATPases belong to the main calcium regulating proteins involved in the balance of calcium homeostasis, which is impaired in oxidative/nitrosative stress and related diseases or aging. The mechanisms of Ca2+-ATPases dysfunction are discussed, focusing on cystein oxidation and tyrosine nitration. Flavonoids act not only as antioxidants but are also able to bind directly to Ca2+-ATPases, thus changing their conformation, which results in modulation of enzyme activity