Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The interaction of Cibacron Blue 3GA (C.I.2) and Remazol Brillant Blue R (C.I.19) with purified preparations of Lactate dehydrogenase (LDH) and (Na+ + K+)-ATPase was studied by means of the enzyme kinetics method. LDH was found to be inhibited by both C.I.2 and C.I.19, with the former being a stronger inhibitor. This may be explained by the fact that in contrast to C.I.19, C.I.2 resembles the whole molecule of the specific cofactor. (Na+ + K+)-ATPase activity was inhibited by both dyes to approximately similar degree. C.I.2 and C.I.19 resemble the ATP molecule to approximately similar extent, particularly as concerns the molecule shape and size. The results obtained confirmed the applicability of C.I.2 and C.I.19 as nucleotide-specific ligands.
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PMID:Interaction of Cibacron Blue 3GA and Remazol Brilliant Blue R with the nucleotide binding site of lactate dehydrogenase and (Na+ + K+)-ATPase. 217 32

Cibacron Blue F3GA (Cb) effectively and reversibly inhibits the activity of (Na,K)-ATPase. Its inhibitory effect does not occur through occupation of the ouabain binding site, but presumably results from Cb-occupation of one catalytic site not competitively attracting ATP. Cb also inhibits ouabain binding to (Na,K)-ATPase. Its inhibitory effect is competitively antagonized by ATP proving accommodation of Cb in the ATP binding site. - If one admits Cb as a suitable analytical tool for the detection of a supersecondary structure folding pattern, the findings suggest that the ATP binding site is lined by beta-pleated sheets flanked by alpha-helices thus providing an environment that funnels ATP to the catalytic site.
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PMID:Kinetic studies on interaction of (Na,K)-ATPase with Cibacron Blue F3GA as probe of the nucleotide fold. 632 87

Cibacron Blue F3GA and its immobilized derivatives have been shown before to bind and inhibit nucleotide-dependent enzymes and, among them, myosin subfragment 1. Experiments have been carried out to examine the mechanism of the subfragment 1--dye interaction. Binding of subfragment 1 to immobilized dye (Affi-Gel Blue) does not involve the ATP binding site on myosin. Subfragment 1 hydrolyzes MgATP and CaATP while bound to the Affi-Gel Blue column. Inactivated subfragment 1, which contains [3H]ADP noncovalently trapped at the active site, binds and elutes from the Affi-Gel Blue column in the same manner as unmodified, active protein. Free Cibacron Blue inhibits the ATPase activity of subfragment 1. The inhibition is pH, salt, and time dependent. Complete inhibition correlates with the noncovalent binding of four to five dye molecules per mole of subfragment 1. Three to four of these dye molecules can be preferentially removed from subfragment 1 in the presence of 1 M KCl without relieving the inhibition. This inhibition, which can be traced to one dye molecule per subfragment 1, is reversible and is facilitated in the presence of MgADP and MgATP, suggesting that the dye does not bind at the active site of subfragment 1. Our observations are explained in terms of hydrophobic and electrostatic protein--dye interactions.
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PMID:Interaction of myosin subfragment 1 with Cibacron Blue F3GA. 645 18

Binding of troponin to Cibacron Blue F3GA-agarose column and its selective release from the gel in the presence of 0.5 M KCl provides the basis for a new purification method. The two-step procedure consists of isoelectric precipitation of tropomyosin and chromatography of the resultant crude troponin supernatant on Affi-Gel Blue column. Adsorption of troponin to the immobilized dye appears to occur through the troponin-T subunit. Troponin-I and troponin-C do not bind to the blue agarose column, whereas troponin-T binds to it very tightly. Binding of the dye to troponin-T prevents formation of troponin-T-troponin-C complex, but does not interfere with direct interaction of troponin-T with troponin-I. The activity of troponin in conferring calcium sensitivity on actomyosin ATPase is not affected by Cibacron Blue. Circular dichroism and difference absorption measurements of complexes of the blue dye with troponin and its subunits reveal the presence of a tight binding site on whole troponin and on troponin-T (KA greater than or equal to 10(6) M). The existence of weak binding sites for the dye on troponin and all of its subunits is deduced from difference absorption studies. Cibacron Blue appears to be a sensitive probe for subunit interactions in troponin.
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PMID:The interaction of Cibacron Blue F3GA with troponin and its subunits. 689 62

A human brain E-type ATPase (HB6 ecto-apyrase) was subjected to site-directed mutagenesis to assess the functional significance of two highly conserved tryptophan residues (Trp 187 and Trp 459), the only two tryptophans conserved in nearly all E-type ATPases. Mutation of tryptophan 187 to alanine yielded a poorly expressed ecto-apyrase completely devoid of nucleotidase activity. Immunolocalization of the W187A mutant in mammalian COS cells showed a cellular distribution clearly different from that of the wild-type enzyme, with the majority of the immunoreactivity concentrated in the interior of the cell. Unlike the wild-type enzyme, this mutant did not bind the nucleotide analogue Cibacron Blue and was sensitive to proteolytic digestion by chymotrypsin. These results suggest alteration of the tertiary structure, causing the enzyme to be improperly folded and retained within the cell. In contrast, mutation of tryptophan 459 to alanine resulted in an ecto-apyrase with enhanced NTPase activity, but diminished NDPase activity. Immunolocalization of this active mutant ecto-apyrase revealed a cellular pattern similar to that of the wild-type enzyme, distributed along the cell periphery and in cell processes. Coupling this active W459A mutation to a previously described mutation (D219E) resulted in an enzyme which preferentially hydrolyzes nucleoside triphosphates over diphosphates. The D219E/W459A double mutant had an ATPase:ADPase ratio of 11:1 and a UTPase:UDPase ratio of 148:1. In addition, the double mutant is substantially less sensitive to inhibition by azide, a more potent inhibitor of ecto-apyrases than ecto-ATPases. Thus, mutation of only two amino acids of an E-type ATPase essentially converts an ecto-apyrase to an ecto-NTPase.
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PMID:Mutagenesis of two conserved tryptophan residues of the E-type ATPases: inactivation and conversion of an ecto-apyrase to an ecto-NTPase. 1023 36

Chicken muscle ecto-ATPase has unusual enzyme kinetics and properties not found in many other E-type ATPases. To determine whether the unique properties of the chicken ecto-ATPase are inherent in the protein sequence and not mediated by some unique property of the chicken system, we have spliced together two partial cDNAs encoding the ecto-ATPase. The enzymatic properties of the COS (green monkey kidney) cell-expressed protein are indistinguishable from the purified chicken gizzard ecto-ATPase, including a 2- to 3-fold stimulation of membrane-bound activity by crosslinking and lectins, properties not shared by most other E-type ATPases. The expressed enzyme is specific for nucleotide triphosphates (ATPase:ADPase hydrolysis ratio of 26:1) and is inhibited by Cibacron Blue (IC50 = 10 microM). The active, expressed enzyme can be affinity-purified with Cibacron Blue, is relatively resistant to deglycosylation, and is less stable than other E-type ATPases. Expression in the presence of tunicamycin resulted in an inactive, unfolded enzyme.
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PMID:Expression and characterization of chicken muscle ecto-ATPase in mammalian COS cells. 1079 17

A specific feature of anthraquinone dyes (AD) is to mimic the adenine nucleotides ATP, ADP, NAD and NADH, enabling them to act as ligands in interaction with nucleotide-binding sites of several enzymes and receptors. In the present study, the interactions and/or inhibitory effects of eight AD, including Cibacron Blue 3G-A (Reactive Blue 2), Procion Blue MX-R (Reactive Blue 4) and Remazol Brilliant Blue R (Reactive Blue 19) on the activity of (Na(+)/K(+))-ATPase were investigated. The AD used in this paper could be divided into two groups: i) AD1-AD4 that do not contain the triazine moiety; ii) AD5-AD8 that contain the triazine moiety. Interaction affinity between the respective dye and (Na+/K+)-ATPase was characterized by means of enzyme kinetics. All AD, excluding AD1 and AD2 (which were practically ineffective) exerted effective competitive inhibition to the (Na(+)/K(+))-ATPase activity. Present study is devoted to elucidation of relationship between the inhibitory efficacy of AD against (Na(+)/K(+))-ATPase activity, their acid-basic properties and their three dimensional structure. From the results obtained, the following conclusions could be driven: 1. Similarities in the mutual position of positively and negatively charged parts of ATP and AD are responsible for their interaction with ATP-binding site of (Na(+)/K(+))-ATPase. This may be documented by fact that mutual position of 1-aminogroup of anthraquinone and -SO3(-) group of benzenesulphonate part of respective AD plays crucial role for inhibition of this enzyme. Distances of these two groups on all effective AD were found to be similar as the distance of the 6-aminogroup of adenine and the second phosphate group on ATP molecule. This similarity could be responsible for biomimetic recognition of AD in ATP-binding loci of (Na(+)/K(+))-ATPase. 2. The affinity of AD to ATP binding site of (Na(+)/K(+))-ATPase increases with increasing values of molar refractivity, i. e., with increasing molecular volume and polarizability.
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PMID:Inhibition of (Na(+)/K(+))-ATPase by Cibacron Blue 3G-A and its analogues. 1735 35