Excitation energy transfer studies on the proximity between SH1 and the adenosinetriphosphatase site in myosin subfragment 1.
AuthorsTao T, Lamkin M
JournalBiochemistry
PubMed ID6457630
'Excitation energy transfer studies were carried out to determine the distance between the adenosinetriphosphatase (ATPase) site and a unique "fast-reacting" sulfhydryl (referred to as SH1) in myosin subfragment 1. The fluorescent moiety of the probe N-(iodoacetyl)-N''-(5-sulfo-1-naphthyl)ethylene-diamine was used as the donor attached at SH1. The chromophoric nucleotide analogue 2''(3'')-0-(2,4,6-trinitrophenyl)adenosine 5''-diphosphate ... More
Trinitrophenyl-ATP and -ADP bind to a single nucleotide site on isolated beta-subunit of Escherichia coli F1-ATPase. In vitro assembly of F1-subunits requires occupancy of the nucleotide-binding site on beta-subunit by nucleoside triphosphate.
AuthorsRao R, Al-Shawi MK, Senior AE
JournalJ Biol Chem
PubMed ID2895769
'The stoichiometry of nucleotide binding to the isolated alpha- and beta-subunits of Escherichia coli F1-ATPase was investigated using two experimental techniques: (a) titration with fluorescent trinitrophenyl (TNP) derivatives of AMP, ADP, and ATP and (b) the centrifuge column procedure using the particular conditions of Khananshvili and Gromet-Elhanan (Khananshvili, D., and ... More
Assessment of the number of nucleotide binding sites on chloroplast coupling factor 1 by the continuous variation method.
AuthorsMusier KM, Hammes GG
JournalBiochemistry
PubMed ID2904277
'The method of continuous variation (Job plot analysis) and difference absorbance spectroscopy were used to investigate the binding of 2''(3'')-(trinitrophenyl)-ADP and -ATP to chloroplast coupling factor 1 (CF1). Experiments performed at a low total concentration (30 microM) of nucleotide and enzyme binding sites (assuming three or four binding sites per ... More
Inhibition study of ADP,ATP transport in mitochondria with trinitrophenyl-modified substrates.
AuthorsSchlimme E, Boos KS, Onur G, Ponse G
JournalFEBS Lett
PubMed ID6301884
'The ADP,ATP carrier of rat liver mitochondria is specifically inhibited by Meisenheimer-type trinitrophenyl (TNP) derivatives of ADP and ATP. Due to a systematic inhibition study we could show that the TNP-moiety itself, even in the 2'', 3''-O-cyclic Meisenheimer complex, revealed no inhibition of mitochondrial ADP,ATP transport. Nucleosidic TNP-compounds are weak ... More
TNP-AMP binding to the sarcoplasmic reticulum Ca(2+)-ATPase studied by infrared spectroscopy.
AuthorsLiu M, Barth A
JournalBiophys J
PubMed ID14581226
'Infrared spectroscopy was used to monitor the conformational change of 2'',3''-O-(2,4,6-trinitrophenyl)adenosine 5''-monophosphate (TNP-AMP) binding to the sarcoplasmic reticulum Ca(2+)-ATPase. TNP-AMP binding was observed in a competition experiment: TNP-AMP is initially bound to the ATPase but is then replaced by beta,gamma-iminoadenosine 5''-triphosphate (AMPPNP) after AMPPNP release from P(3)-1-(2-nitrophenyl)ethyl AMPPNP (caged AMPPNP). ... More
2', 3'-O-(2,4,6,trinitrophenyl)-ATP and A-317491 are competitive antagonists at a slowly desensitizing chimeric human P2X3 receptor.
'(1) Rapid desensitization of ligand-gated ion channel receptors can alter the apparent activity of receptor modulators, as well as make detection of fast-channel activation difficult. Investigation of the antagonist pharmacology of ATP-sensitive homomeric P2X3 receptors is limited by agonist-evoked fast-desensitization kinetics. (2) In the present studies, chimeric receptors were created ... More
2,4,6-Trinitrobenzenesulfonate labels an essential amino group near the bound phosphate at the catalytic site of mitochondrial F1-ATPase.
AuthorsTing LP, Wang JH
JournalBiochem Biophys Res Commun
PubMed ID6458293
Fluorescence changes of a label attached near the myosin active site on nucleotide binding in rat skeletal muscle fibres.
AuthorsFujita S, Nawata T, Yamada K
JournalJ Physiol
PubMed ID10066911
'1. Trinitrophenyl AMP (TNP-AMP) in the concentration range 10-300 microM induced an increase in fluorescence intensity at around 530 nm in skinned skeletal muscle fibres freshly obtained from rat psoas muscle. 2. The fluorescence intensity of the fibres depended on TNP-AMP concentration up to approximately 200 microM. The Kd of ... More
Fluorescent and colored trinitrophenylated analogs of ATP and GTP.
AuthorsHiratsuka T
JournalEur J Biochem
PubMed ID12919312
Fluorescent and colored trinitrophenylated (TNP) analogs of ATP and GTP can interact with nucleotide-requiring enzymes and proteins as a substitute for the parent nucleotide. These analogs have strong binding affinities for most nucleotide-requiring systems. Their bindings are easily detected by absorption and fluorescence changes in the visible region. Recent years ... More
Using 2'(3')-O-trinitrophenyl derivatives of adenine nucleotides to study the structure and mechanism of functioning of soluble mitochondrial ATPase.
The 2'(3')-O-trinitrophenyl (N3ph) derivatives of the adenine nucleotides are strong competitive inhibitors of isolated mitochondrial ATPase (factor F1). Ki decreases in the order N3phAdo greater than N3phAdo greater than N3phAMP greater than N3phADP and is equal to 8 nM for N3phADP. Picric acid, which activates the ATPase reaction of factor ... More
Adenosine 5'-monophosphate inhibits the association of 14-3-3 proteins with the plant plasma membrane H(+)-ATPase.
AuthorsCamoni L, Visconti S, Marra M, Aducci P
JournalJ Biol Chem
PubMed ID11423544
Although a well ascertained evidence proves that the activity of the plant plasma membrane H(+)-ATPase is regulated by 14-3-3 proteins, information about physiological factors modulating the phosphorylation-dependent association between 14-3-3 proteins and the proton pump is largely incomplete. In this paper we show that the 5'-AMP-mimetic, 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), inhibits ... More
Biological activities and spectroscopic properties of chromophoric and fluorescent analogs of adenine nucleoside and nucleotides, 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) adenosine derivatives.
AuthorsHiratsuka T
JournalBiochim Biophys Acta
PubMed ID6295507
The ribose-modified chromophoric and fluorescent analog of ATP 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) adenosine 5'-triphosphate (TNP-ATP) has been synthesized previously (Hiratsuka, T., and Uchida, K. (1973) Biochim. Biophys. Acta 320, 635-647 and Hiratsuka, T. (1976) Biochim. Biophys. Acta 453, 293-297). In the present study, four TNP-derivatives of ATP, ADP, AMP and adenosine were synthesized ... More
Expression and functional properties of the second predicted nucleotide binding fold of the cystic fibrosis transmembrane conductance regulator fused to glutathione-S-transferase.
AuthorsRandak C, Roscher AA, Hadorn HB, Assfalg-Machleidt I, Auerswald EA, Machleidt W
JournalFEBS Lett
PubMed ID7537226
CFTR-NBF-2 was expressed in Escherichia coli in fusion with glutathione-S-transferase, the soluble portion was purified and identified as a structured protein by its CD spectrum. Association reactions of the recombinant NBF-2 with adenine nucleotides were monitored qualitatively by demonstrating its ability to bind specifically to ATP-, ADP- and AMP-affinity agarose ... More
Cystic fibrosis transmembrane conductance regulator: nucleotide binding to a synthetic peptide.
Multiple mutations in the gene responsible for cystic fibrosis are located within a region predicted to encode a nucleotide-binding fold in the amino terminal half of the cystic fibrosis transmembrane conductance regulator protein. A 67-amino acid peptide (P-67) that corresponds to the central region of this putative nucleotide binding site ... More
Inhibition of sarcoplasmic reticulum Ca2+-ATPase by Mg2+ at high pH.
AuthorsBishop JE, Al-Shawi MK
JournalJ Biol Chem
PubMed ID2962998
Steady state turnover of Ca2+-ATPase of sarcoplasmic reticulum has generally been reported to have a bell-shaped pH profile, with an optimum near pH 7.0. While a free [Mg2+] of 2 mM is optimal for activity at pH 7.0, it was found that this level was markedly inhibitory (K1/2 = 2 ... More
Oxidation of thiols in the Ca2+-ATPase of sarcoplasmic reticulum microsomes.
AuthorsScherer NM, Deamer DW
JournalBiochim Biophys Acta
PubMed ID2946320
We recently showed that oxidative stress impairs the function of the sarcoplasmic reticulum to transport and retain calcium. Inhibition results primarily from oxidation of one or more thiol groups in the Ca2+-ATPase. We now report that thiol oxidation does not result in disulfide formation. Oxidative inhibition of Ca2+-ATPase activity was ... More
Mutations in the hinge of a dynamic loop broadly influence functional properties of fructose-1,6-bisphosphatase.
AuthorsNelson SW, Choe JY, Honzatko RB, Fromm HJ
JournalJ Biol Chem
PubMed ID10896931
Loop 52-72 of porcine fructose-1,6-bisphosphatase may play a central role in the mechanism of catalysis and allosteric inhibition by AMP. The loop pivots between different conformational states about a hinge located at residues 50 and 51. The insertion of proline separately at positions 50 and 51 reduces k(cat) by up ... More
Glucose phosphorylation. Interaction of a 50-amino acid peptide of yeast hexokinase with trinitrophenyl ATP.
AuthorsArora KK, Shenbagamurthi P, Fanciulli M, Pedersen PL
JournalJ Biol Chem
PubMed ID2318895
A 50-amino acid peptide predicted by chemical modification studies of yeast hexokinase to contain an ATP-binding site has been synthesized and purified. The peptide, which includes residues from glutamate 78 at the NH2-terminal end to leucine 127 at the COOH-terminal, resides within the smaller of the two lobes found in ... More
The presence of two hydrolytic sites on beef heart mitochondrial adenosine triphosphatase.
AuthorsGrubmeyer C, Penefsky HS
JournalJ Biol Chem
PubMed ID6452454
The ribose-modified nucleotides 2',3'-O-(2,4,6-trinitrophenyl) adenosine 5'-triphosphate (TNP-ATP) and TNP-ADP were used to probe the catalytic sites on soluble beef heart mitochondrial adenosine triphosphatase (F1). Both compounds were potent competitive inhibitors of ATP hydrolysis catalyzed by F1, Ki = 5.5 and 10 nM, respectively, and by submitochondrial particles, Ki (TNP-ATP) = ... More
Structural requirements for the binding of AMP to the allosteric site of NAD-specific isocitrate dehydrogenase from bakers' yeast.
AuthorsGabriel JL, Plaut GW
JournalBiochemistry
PubMed ID2162196
The specificity of yeast NAD-specific isocitrate dehydrogenase for the structures of the allosteric effector 5'-AMP was examined with analogues modified in the purine ring, pentosyl group, and 5'-phosphate group. An unsubstituted 6-amino group was essential for activation as was the phosphoryl group at the 5'-position. Activity was retained when an ... More
Two distinct classes of nucleotide binding sites in sarcoplasmic reticulum Ca-ATPase revealed by 2',3'-O-(2,4,6-trinitrocyclohexadienylidene)-ATP.
AuthorsDupont Y, Pougeois R, Ronjat M, Verjovsky-Almeida S
JournalJ Biol Chem
PubMed ID3158655
It was previously reported that 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) (TNP)-nucleotides bind with high affinity to the sarcoplasmic reticulum Ca-ATPase (Dupont, Y., Chapron, Y., and Pougeois, R. (1982) Biochem. Biophys. Res. Commun. 106, 1272-1279 and Watanabe, T., and Inesi, G. (1982) J. Biol. Chem. 257, 11510-11516). Here we report a study of the Ca-ATPase ... More
Structural dynamics of the Ca2(+)-ATPase of sarcoplasmic reticulum. Temperature profiles of fluorescence polarization and intramolecular energy transfer.
AuthorsJona I, Matko J, Martonosi A
JournalBiochim Biophys Acta
PubMed ID2145977
The temperature dependence of fluorescence polarization and Förster-type resonance energy transfer (FRET) was analyzed in the Ca2(+)-ATPase of sarcoplasmic reticulum using protein tryptophan and site-specific fluorescence indicators such as 5-[2-[iodoacetyl)amino)ethyl]aminonaphthalene-1-sulfonic acid (IAEDANS), fluorescein 5'-isothiocyanate (FITC), 2',3'-O-(2,4,3-trinitrophenyl)adenosine monophosphate (TNP-AMP) or lanthanides (Pr3+, Nd3+) as probes. The normalized energy transfer efficiency between ... More
Kinetic regulation of yeast NAD-specific isocitrate dehydrogenase by citrate.
AuthorsGabriel JL, Plaut GW
JournalBiochemistry
PubMed ID2001349
The present results suggest that the enzyme modifier citrate and the substrate isocitrate are bound at different sites on yeast NAD-specific isocitrate dehydrogenase and that citrate diminishes the binding of the positive effector 5'-AMP, thereby causing a decreased rate of enzyme catalysis. This interpretation differs from the earlier proposal that ... More
Inhibition of sodium and potassium adenosine triphosphatase by 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) adenine nucleotides. Implications for the structure and mechanism of the Na:K pump.
AuthorsMoczydlowski EG, Fortes PA
JournalJ Biol Chem
PubMed ID6257716
Trinitrophenyl derivatives of adenine nucleotides (TNP-nucleotides: 2',3'-O-2,4,6-trinitrocyclohexadienylidene complexes at neutral or basic pH) are potent inhibitors of (Na,K)-ATPase activity. The inhibitory potency of the derivatives tested followed the sequence: TNP-ADP greater than TNP-ATP greater than TNP-AMP much greater than TNP-IMP greater than TNP-adenosine. In the presence of Na+ plus K+, ... More
Mutational analysis of the consensus nucleotide binding sequences in the rat liver mitochondrial ATP synthase beta-subunit.
AuthorsThomas PJ, Garboczi DN, Pedersen PL
JournalJ Biol Chem
PubMed ID1400352
The coupling step in the biosynthesis of ATP in biological systems is generally believed to involve an energy-requiring release of ATP bound to the beta-subunit of the ATP synthase complex. A molecular description of the ATP binding site on the beta-subunit is, therefore, critical to understanding the mechanism of coupling ... More
Negative cooperativity in the binding of nucleotides to Escherichia coli replicative helicase DnaB protein. Interactions with fluorescent nucleotide analogs.
AuthorsBujalowski W, Klonowska MM
JournalBiochemistry
PubMed ID8504109
The interactions of nucleotides with Escherichia coli replicative helicase DnaB protein have been systematically studied using fluorescent nucleotide analogs, 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate (TNP-ATP), 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-diphosphate (TNP-ADP), 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-monophosphate (TNP-AMP), 3'-O-(N-methylantraniloyl) 5'-diphosphate (MANT-ADP), and 1,N6-ethenoadenosine diphosphate (epsilon ADP). The binding of the analogs is accompanied by strong quenching of the protein fluorescence; ... More
2',3'-O-(2,4,6-trinitrophenyl)-8-azido-adenosine mono-, di-, and triphosphates as photoaffinity probes of the Ca2+-ATPase of sarcoplasmic reticulum. Regulatory/superfluorescent nucleotides label the catalytic site with high efficiency.
AuthorsSeebregts CJ, McIntosh DB
JournalJ Biol Chem
PubMed ID2521624
We have synthesized a new class of ATP photo-affinity analogs, 2',3'-O-(2,4,6-trinitrophenyl)-8-azido (TNP-8N3)-ATP, -ADP, and -AMP, and their radiolabeled derivatives, and characterized their interaction with sarcoplasmic reticulum vesicles. The nucleotides bind with high affinity (Kd = 0.04-0.4 microM) to the catalytic site of the Ca2+-ATPase. TNP-8N3-ATP and TNP-8N3-ADP, at low concentrations ... More
Cooperatively between catalytic sites in the mechanism of action of beef heart mitochondrial adenosine triphosphatase.
AuthorsGrubmeyer C, Penefsky HS
JournalJ Biol Chem
PubMed ID6452455
Occupancy of only one of two hydrolytic sites on beef heart mitochondrial ATPase (F1) by the radioactive ATP analog, 2',3'-O-(2,4,6-trinitrophenyl) adenosine 5'-[gamma-32P]-triphosphate (TNP-[gamma-32P]ATP) is associated with a low rate of hydrolysis of the substrate even under conditions otherwise favoring catalysis. Addition of excess nonradioactive TNP-ATP, in concentrations sufficient to fill ... More
ATP regulation of sarcoplasmic reticulum Ca2+-ATPase. Metal-free ATP and 8-bromo-ATP bind with high affinity to the catalytic site of phosphorylated ATPase and accelerate dephosphorylation.
To localize and characterize the regulatory nucleotide site of skeletal muscle sarcoplasmic reticulum Ca2+-ATPase, we have investigated the effects of ADP, ATP, and analogues of these nucleotides on the rate of dephosphorylation of both native ATPase and ATPase modified with fluorescein 5'-isothiocyanate (FITC), a reagent which hinders access of nucleotides ... More
The effect of high pressure on the conformation, interactions and activity of the Ca(2+)-ATPase of sarcoplasmic reticulum.
AuthorsJona I, Martonosi A
JournalBiochim Biophys Acta
PubMed ID1837234
High pressure (100-150 MPa) increases the intensity and polarization of fluorescence of FITC-labeled Ca(2+)-ATPase in a medium containing 0.1 mM Ca2+, suggesting a reversible pressure-induced transition from the E1 into an E2-like state with dissociation of ATPase oligomers. Under similar conditions but using unlabeled sarcoplasmic reticulum vesicles, high pressure caused ... More