140 research outputs found
Effects of reactive oxygen and nitrogen species on actomyosin and their implications for muscle contractility
Experimental evidence accumulated during recent years is pointing out that numerous pathological conditions in skeletal and cardiac muscle are associated with an oxidative stress-induced muscle injury. Additionally, it has been postulated that several oxidants can directly alter contractile function by oxidative modification of the myofibril proteins –
actin and myosin. Peroxynitrite (ONOO-), a potent biological oxidizing agent formed in the nearly instantaneous reaction of nitric oxide with superoxide anion, is increasingly recognized as playing a major role in the skeletal and cardiac muscle dysfunction. This is supported by detection of 3-nitrotyrosine, a protein modification produced by the reaction of peroxynitrite with tyrosine, on skeletal and cardiac muscle proteins during aging or in diseases associated with myocardial inflammation or ischemia/reperfusion insults. Although some studies point to a correlation of protein nitration with functional and structural modifications, the mechanism by which peroxynitrite may impair muscle contractility remains far from being elucidated. In the present review we address the role of reactive oxygen and nitrogen species on the structural and functional impairment of actomyosin ATPase activity and their implications for muscle contraction with particular emphasis on the oxidative modifications promoted by peroxynitrite on actin and myosin
Estudios químico-físicos de la interacción de efectores alostéricos con fosforilasa b de músculo de esqueleto de conejo
La fosforilasa b es una enzima alostérica ampliamente estudiada. Su activación requiere la presencia de amp siendo sus inhibidores alostéricos el atp y la glucosa-6-fosfato. Aunque comenzó a estudiarse en la década de los 30 quedan aun muchos problemas por aclarar. Para intentar resolver estos problemas se han utilizado en el presente estudio los resultados obtenidos por: a) activación e inhibición de la enzima en presencia de análogos de amp en la parte básica ribosa y fosfato. B) cinéticas de valoración de sh con dtnb. C) medidas de ultracentrifugación d) medidas calorimétricas en función de t y concentración de la enzima
Monomeric versus decameric vanadate in the elucidation of muscle contraction regulation: a kinetic, spectroscopic and structural overview
Vanadium (V) was rediscovered for biology as a “muscle inhibitor factor” when it was found in commercial ATP prepared from equine muscle almost thirty years ago. Since
then it has been used as a molecular probe of the mechanisms of several enzyme reactions involving hydrolysis of phosphate
ester bonds. Besides acting as a phosphate analogue, vanadate has also the potential to exhibit biological activities through oligomeric vanadate species. Among the vanadate oligomers, decavanadate is one of the most potent inhibitors and has revealed an excellent kinetic and spectroscopic
probe. This is particularly relevant for myosin, the major muscle ATPase which along with actin is able to convert the
chemical energy of ATP hydrolysis into mechanical work.
Apparently, vanadate is able to populate different conformational states of the myosin ATPase cycle depending
on its oligomerization state. While monomeric vanadate (VO4
3-) mimics the transition state for the g-phosphate hydrolysis blocking myosin in a pre-power stroke state, decameric vanadate (V10O28 6-) induces the formation of the
intermediate myosin·MgATP·V10 complex blocking the actomyosin cycle in a pre-hydrolysis state. These recent
findings, that are now reviewed, point out to the importance of taking into account vanadate species variety in studies
describing the interaction of vanadate with biological systems and incite the use of decavanadate as a biochemical tool to the elucidation of muscle contraction regulation
Estudios químico-físicos de la interacción de efectores alostéricos con fosforilasa b de músculo de esqueleto de conejo
Tesis inédita de la Universidad Complutense de Madrid, Facultad de Ciencias Químicas, leída el 09-07-1977.La fosforilasa b es una enzima alostérica ampliamente estudiada. Su activación requiere la presencia de amp siendo sus inhibidores alostéricos el atp y la glucosa-6-fosfato. Aunque comenzó a estudiarse en la década de los 30 quedan aun muchos problemas por aclarar. Para intentar resolver estos problemas se han utilizado en el presente estudio los resultados obtenidos por: a) activación e inhibición de la enzima en presencia de análogos de amp en la parte básica ribosa y fosfato. B) cinéticas de valoración de sh con dtnb. C) medidas de ultracentrifugación d) medidas calorimétricas en función de t y concentración de la enzima.Fac. de Ciencias QuímicasTRUEProQuestpu
John's ellipsoid and the integral ratio of a log-concave function
We extend the notion of John’s ellipsoid to the setting of integrable
log-concave functions. This will allow us to define the integral ratio of a
log-concave function, which will extend the notion of volume ratio, and we
will find the log-concave function maximizing the integral ratio. A reverse
functional affine isoperimetric inequality will be given, written in terms of this
integral ratio. This can be viewed as a stability version of the functional affine
isoperimetric inequality.Ministerio de Economía y CompetitividadFondo Europeo de Desarrollo RegionalConsejería de Industria, Turismo, Empresa e Innovación (Comunidad Autónoma de la Región de Murcia)Coordenação de aperfeiçoamento de pessoal de nivel superiorInstituto Nacional de Matemática Pura e Aplicad
Decavanadate induces mitochondrial membrane depolarization and inhibits oxygen consumption
Decavanadate induced rat liver mitochondrial depolarization at very low concentrations, half-depolarization with 39 nM decavanadate,
while it was needed a 130-fold higher concentration of monomeric vanadate (5 lM) to induce the same effect. Decavanadate also
inhibits mitochondrial repolarization induced by reduced glutathione in vitro, with an inhibition constant of 1 lM, whereas no effect was observed up to 100 lM of monomeric vanadate. The oxygen consumption by mitochondria is also inhibited by lower decavanadate than monomeric vanadate concentrations, i.e. 50% inhibition is attained with 99 nM decavanadate and 10 lM monomeric vanadate. Thus,
decavanadate is stronger as mitochondrial depolarization agent than as inhibitor of mitochondrial oxygen consumption. Up to 5 lM,
decavanadate does not alter mitochondrial NADH levels nor inhibit neither FOF1-ATPase nor cytochrome c oxidase activity, but it
induces changes in the redox steady-state of mitochondrial b-type cytochromes (complex III). NMR spectra showed that decameric vanadate is the predominant vanadate species in decavanadate solutions. It is concluded that decavanadate is much more potent mitochondrial depolarization agent and a more potent inhibitor of mitochondrial oxygen consumption than monomeric vanadate, pointing out the importance to take into account the contribution of higher oligomeric species of vanadium for the biological effects of vanadate solutions
Decavanadate toxicity effects following in vivo administration
Very few in vivo animal studies involving vanadium consider the contribution of decavanadate (V10) to vanadium
biological effects. Recently, it is been suggested that decameric vanadate may not completely fall apart into other vanadate oligomers before induces changes in cell
homeostasis, namely in several stress markers. An acute exposure of different fish species (Halobactrachus didactilus,
Lusitanian toadfish, and Sparus aurata, gilthead seabream) to decavanadate, but not to other vanadate oligomers,
induced different effects than vanadate in catalase activity, glutathione content, lipid peroxidation, mitochondrial
superoxide anion production and vanadium accumulation, whereas both solutions seem to equally depress reactive oxygen species (ROS) production as well as total
intracellular reducing power. Vanadium is accumulated in Sparus aurata mitochondria in particular when decavanadate is administrated. Moreover, exposure to different vanadate oligomers induced morphological changes in fish cardiac,
hepatic and renal tissues causing tissues lesions in the liver and kidney, but not cardiac tissue. Nevertheless, the results
highlight that different vanadate oligomers seem to follow, not only in vitro but also in vivo, different pathways, with different targets and effects. These recent findings, that are now summarized, point out the decameric vanadate species contributions to in vivo effects induced by vanadium in
biological systems
Binding modes of decavanadate to myosin and inhibition of the actomyosin ATPase activity
Decavanadate, a vanadate oligomer, is known to interact with myosin and to inhibit the ATPase activity, but the putative binding sites and the mechanism of inhibition are still to be clarified. We have previously proposed that the decavanadate (V10O28
6−) inhibition of the actin-stimulated
myosin ATPase activity is non-competitive towards both actin and ATP. A likely explanation for these results is that V10 binds to the so-called back-door at the end of the Pi-tube opposite to the nucleotide-binding site. In order to further investigate this possibility, we have carried out molecular docking simulations of the V10 oligomer on three different structures of the myosin motor domain of Dictyostelium discoideum, representing distinct states of the ATPase cycle. The results indicate a clear preference of V10 to bind at the back-door, but only on the “open”
structures where there is access to the phosphate binding-loop. It is suggested that V10 acts as a “back-door stop” blocking the closure of the 50-
kDa cleft necessary to carry out ATP-γ-phosphate hydrolysis. This provides a simple explanation to the non-competitive behavior of V10 and spurs the use of the oligomer as a tool to elucidate myosin back-door conformational changes in the process of muscle contraction
Peroxynitrite induces F-actin depolymerization and blockade of myosin ATPase stimulation
Treatment of F-actin with the peroxynitrite-releasing agent 3-morpholinosydnonimine (SIN-1) produced a dose-dependent F-actin
depolymerization. This is due to released peroxynitrite because it is not produced by ‘decomposed SIN-1’, and it is prevented by
superoxide dismutase concentrations efficiently preventing peroxynitrite formation. F-actin depolymerization has been found to be very sensitive to peroxynitrite, as exposure to fluxes as low as 50–100 nM peroxynitrite leads to nearly 50% depolymerization in about 1 h.
G-actin polymerization is also impaired by peroxynitrite although with nearly 2-fold lower sensitivity. Exposure of F-actin to submicromolar fluxes of peroxynitrite produced cysteine oxidation and also a blockade of the ability of actin to stimulate myosin ATPase activity.
Our results suggest that an imbalance of the F-actin/G-actin equilibrium can account for the observed structural and functional impairment of myofibrils under the peroxynitrite-mediated oxidative stress reported for some pathophysiological conditions
The pathways of cell death in cardiomyocytes induced by vanadate
After 24 hours, cardiac myocytes exposure to 10 μM (LD50) vanadate (meta or decavanadate) an increased (30%) of caspase 3-activation was observed, although not significant. On contrary, a significant decrease (40%) of
ATP content, characteristic of necrotic cell death was detected. Furthermore,
vanadate treatment increased intracellular Ca2+ level from 60 nM to 240 nM,
whereas it decreases mitochondria superoxide anion generation and induces
mitochondria membrane depolarization (IC50=6.5 μM). In conclusion, micromolar
vanadate exposure induces large chances in two major bioenergetic markers in cardiac myocytes: intracellular calcium concentration and superoxide anion
mitochondrial production, suggesting a necrotic cell death through a mitochondrial
toxic pathway
- …