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Vanadate
About: Vanadate is a research topic. Over the lifetime, 4497 publications have been published within this topic receiving 120109 citations. The topic is also known as: vanadate.
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TL;DR: Results suggest that the Neurospora ATPase undergoes significant conformational changes at both termini of the polypeptide during its reaction cycle.
42 citations
TL;DR: ATPase transport in reconstituted vesicles was similar to that in native vesicle with respect to substrate specificity, salt stimulation, and inhibitor sensitivity, indicating a relatively nonspecific interaction of the H+- ATPase with the hydrophobic lipid environment.
Abstract: The distribution of corn root microsomal ATPase activity on continuous sucrose gradients was nearly identical to that of membrane vesicles capable of ATP-dependent H+ transport when assayed in the presence of 20 μm vanadate, 3mm azide, 5mm MgSO4 and 50mm LiCl. These assay conditions were used to monitor solubilization of an anion-sensitive H+-ATPase from a purified membrane fraction (20–30% (wt/wt) sucrose interface) believed to be derived from tonoplast membranes.
42 citations
TL;DR: TheVanadate-induced inhibition of 30S dynein Mg-ATPase was noncompetitive in the entire range of ATP concentration examined, and the dissociation constant of vanadate and the molecular weight per enzymatic active site according to the kinetics of tight-binding inhibition with several assumptions was estimated.
Abstract: Inhibitory action of vanadate (orthovanadate and metavanadate) on ciliary dynein adenosinetriphosphatase (ATPase) from Tetrahymena was investigated. The apparent concentrations of vanadate giving half-maximal inhibition of Mg-ATPase activity of various dynein fractions were as follows: the axoneme-bound form of dynein at 100 nM, solubilized crude dynein at 50 nM, 14S dynein at 5 microM, and 30S dynein at 20 nM. The Ca-ATPase of 30S dynein was more than 30-fold less sensitive than its Mg-ATPase, and still less sensitive was the Ca-ATPase of 14S dynein. The Mg-ATPase of 30S dynein was most sensitive to vanadate at neutral pH, and the addition of KCl or NaCl into the assay mixture reduced its sensitivity. Varying the assay temperature between 0 and 37 degrees C affected the sensitivity to a slight extent. Metavanadate was as much a potent inhibitor of dynein ATPase as orthovanadate, but vanadium pentoxide was less potent. When the dynein ATPase activity was reciprocally plotted against the concentration of vanadate (the Dixon plot), the inhibition was proved to be biphasic. At lower concentrations of vanadate, the inhibition was more significant. Therefore the Dixon plot had a downward bent. Reexamination of the Lineweaver-Burk plot of 30S dynein Mg-ATP showed a downward bent, which indicates that 30S dynein may have at least two Km values, ca. 1 microM and 3 microM; or otherwise, 30S dynein might possibly have a negatively cooperative nature (Hill coefficient 0.67). The vanadate-induced inhibition of 30S dynein Mg-ATPase was noncompetitive in the entire range of ATP concentration examined. Since the vanadate-induced inhibition of 30S dynein Mg-ATPase could be classified into "tight-binding inhibition", we could estimate the dissociation constant of vanadate and the molecular weight per enzymatic active site according to the kinetics of tight-binding inhibition with several assumptions. Thus, the dissociation constant was 10-15 nM, depending on the ATPase assay condition, while the molecular weight per enzymatic active site was 420 000-480 000, independent of the assay condition with the assumption that the present 30S dynein preparation is totally pure. This value would be reduced about 20% when the purity was taken into consideration.
42 citations
TL;DR: It is demonstrated that osteoclasts have a plasma membrane Ca2+‐ATPase with characteristics similar to the enzyme responsible for active calcium extrusion in other cells.
Abstract: The plasma membrane fraction of chicken osteoclasts was purified utilizing 20% continuous Percoll gradients Biochemical marker enzyme analysis (ouabain-sensitive Na+, K+-ATPase and 5′-nucleotidase) indicated that plasma membrane enrichment was 1187-fold and 725-fold, respectively, and contamination with mitochondria, endoplasmic reticulum, and lysosomes was low as determined by succinic dehydrogenase, NADH dehydrogenase, and N-acetylglucosaminidase activities, respectively SDS latency of Na+K+-ATPase and 5′-nucleotidase activities of the isolated plasma membranes revealed that 43-50% of vesicles were sealed, with 10-16% in the inside-out orientation, depending on the membrane fraction used Electron microscopy confirmed the vesicular nature of the plasma membrane fraction The plasma membrane Ca2+-ATPase had a high-affinity (KCa = 022 μM; Kmax = 016 μmol/mg per min) and a low-affinity (KCa = 148 μM; Vmax = 037 μtmol/mg per min) component Calmodulin (012 μM) had no effect on Ca2+-ATPase activity However, trifluoperazine (01 mM), a calmodulin antagonist, strongly inhibited especially the high-affinity component of the enzyme Vanadate and lanthanum also caused inhibition In the presence of CDTA, a potent Ca2+ and Mg2+ chelating agent, high-affinity Ca2+-ATPase activity was abolished, indicating that trace Mg2+ was essential for activity The Ca2+-ATPase substrate curve using ATP showed a high-affinity (Km = 123 μM; Kmax= 0022 μmol/mg per min) and a low-affinity (Km = 438 μM; Vmax = 0278 μmol/mg per min) component These results demonstrate that osteoclasts have a plasma membrane Ca2+-ATPase with characteristics similar to the enzyme responsible for active calcium extrusion in other cells
42 citations
TL;DR: The results are consistent with the notion that SPS contains phosphorylation site(s) that reduce enzyme activation state and that dephosphorylation of these residue(S) is the mechanism of light activation.
42 citations