Simultaneous determination of myo-inositol and scyllo-inositol by MEKC as a rapid monitoring tool for inositol levels.
AuthorsTzeng HF, Chen JY, Huang SW, Wang YJ, Yang CS
JournalElectrophoresis
PubMed ID17351885
'A simple and rapid MEKC method was developed for the simultaneous determination of myo-inositol, scyllo-inositol, and glucose. Prior to electrophoretic separation, the nonfluorescent inositols and glucose were derivatized by N-methylisatoic anhydride at 25 degrees C for 10 min so that they could be detected by a fluorescence detector during separation. ... More
Kinetics of Cdc42 membrane extraction by Rho-GDI monitored by real-time fluorescence resonance energy transfer.
AuthorsNomanbhoy TK, Erickson JW, Cerione RA
JournalBiochemistry
PubMed ID10026253
'The mechanisms underlying the ability of the Rho-GDP dissociation inhibitor (RhoGDI) to elicit the release of Rho-related GTP-binding proteins from membranes is currently unknown. In this report, we have set out to address this issue by using fluorescence resonance energy transfer approaches to examine the functional interactions of the RhoGDI ... More
Kinetics of the interaction of 2'(3')-O-(N-methylanthraniloyl)-ATP with myosin subfragment 1 and actomyosin subfragment 1: characterization of two acto-S1-ADP complexes.
AuthorsWoodward SK, Eccleston JF, Geeves MA
JournalBiochemistry
PubMed ID1824820
'We have used a fluorescent analogue of ATP, mantATP [2''(3'')-O-(N-methylanthraniloyl)-adenosine 5''-triphosphate; Hiratsuka T. (1983) Biochim. Biophys. Acta 742, 496-508], and made a detailed kinetic study of the interaction of mantATP and mantADP with S1 and acto-S1. We have shown that these analogues behave like ATP and ADP, respectively. In addition, ... More
Low affinity interactions of GDPbetaS and ribose- or phosphoryl-substituted GTP analogues with the heterotrimeric G protein, transducin.
'We have examined the effects of three commonly used classes of guanine nucleotide analogues on the retinal G protein, transducin (Gt), and found them to be quite different from those that might be expected from results with other GTP-binding proteins. The most surprising results were with guanosine 5''-O-(2-thiodiphosphate) (GDPbetaS); rather ... More
A study of protein dynamics from anisotropy decays obtained by variable frequency phase-modulation fluorometry: internal motions of N-methylanthraniloyl melittin.
AuthorsMaliwal BP, Hermetter A, Lakowicz JR
JournalBiochim Biophys Acta
PubMed ID3756174
'Internal motions of melittin and its lipid complexes were studied by anisotropy decays determined by frequency-domain fluorometry. A covalent anthraniloyl probe was attached, probably to lysine-21. The emission spectra indicate that the anthraniloyl moiety is exposed to solvent in both monomeric and tetrameric forms and is present at the lipid-water ... More
The HPr kinase from Bacillus subtilis is a homo-oligomeric enzyme which exhibits strong positive cooperativity for nucleotide and fructose 1,6-bisphosphate binding.
AuthorsJault JM, Fieulaine S, Nessler S, Gonzalo P, Di Pietro A, Deutscher J, Galinier A
JournalJ Biol Chem
PubMed ID10636874
'Carbon catabolite repression allows bacteria to rapidly alter the expression of catabolic genes in response to the availability of metabolizable carbon sources. In Bacillus subtilis, this phenomenon is controlled by the HPr kinase (HprK) that catalyzes ATP-dependent phosphorylation of either HPr (histidine containing protein) or Crh (catabolite repression HPr) on ... More
Fluorescence resonance energy transfer on the voltage-dependent sodium channel. Spatial relationship and site coupling between the batrachotoxin and Leiurus quinquestriatus quinquestriatus alpha-scorpion toxin receptors.
AuthorsAngelides KJ, Brown GB
JournalJ Biol Chem
PubMed ID6327667
'A fluorescent N- methylanthraniloyl derivative of the potent depolarizing agent batrachotoxin has been used to probe the structural and conformational properties of the neurotoxin receptor site on the voltage-dependent sodium channel. Batrachotoxin A 20-alpha-N- methylanthranilate (BTX-NMA) retains high affinity for its receptor site on the synaptosomal sodium channel with a ... More
Fluorometric titration of the mitochondrial ADP/ATP carrier protein in muscle homogenate with atractyloside derivatives.
AuthorsRoux P, Le Saux A, Fiore C, Schwimmer C, Dianoux AC, Trézéguet V, Vignais PV, Lauquin GJ, Brandolin G
JournalAnal Biochem
PubMed ID8742079
'We describe here the chemical synthesis of the novel methylanthraniloyl (Mant-) derivative of atractyloside (ATR), which is a specific inhibitor of the mitochondrial ADP/ATP carrier. The spectral properties of Mant-ATR and naphthoyl-ATR (N-ATR) are analyzed. Both derivatives bind to the membrane-bound ADP/ATP carrier at the same sites as ATR and ... More
Polysaccharide labeling with N-methylisatoic anhydride: generation of ultraviolet chromophores and blue fluorophores.
AuthorsDeAngelis PL
JournalAnal Biochem
PubMed ID10933873
Interaction of a fluorescent procaine analogue with phosphatidylcholine vesicles.
AuthorsTrzos W, Reed JK
JournalFEBS Lett
PubMed ID7238879
Covalent attachment of fluorescent groups to the 5'-end of transfer RNA.
AuthorsYang CH, Söll D
JournalArch Biochem Biophys
PubMed ID4351348
Cyclization under mild conditions of anthraniloyl- and N-methyl-anthraniloyl dipeptides.
AuthorsPinnen F, Di Muro A, Zanotti G, Lucente G
JournalInt J Pept Protein Res
PubMed ID3692685
Anthraniloyl- and N-methyl-anthraniloyl dipeptides containing C-terminal L and D proline residues have been synthesized and subjected to cyclization reaction under mild conditions. The influence of substituents and of the chirality of the residues on the output of the reaction is discussed. ... More
The novel fluorescent CDP-analogue (Pbeta)MABA-CDP is a specific probe for the NMP binding site of UMP/CMP kinase.
AuthorsRudolph MG, Veit TJ, Reinstein J
JournalProtein Sci
PubMed ID10631985
Direct thermodynamic and kinetic investigations of the binding of nucleotides to the nucleoside monophosphate (NMP) site of NMP kinases have not been possible so far because a spectroscopic probe was not available. By coupling a fluorescent N-methylanthraniloyl- (mant) group to the beta-phosphate of CDP via a butyl linker, a CDP ... More
The aggregation state of mellitin in lipid bilayers. An energy transfer study.
AuthorsHermetter A, Lakowicz JR
JournalJ Biol Chem
PubMed ID3722154
We used fluorescence non-radiative energy transfer to measure the self-association of melittin in solution and when bound to lipid bilayers. Energy transfer occurred from the tryptophan residue of unlabeled melittin to an N-methyl anthraniloyl residue covalently bound to a basic lysine residue on melittin. The extent of energy transfer from ... More
Batrachotoxinin-A N-methylanthranilate, a new fluorescent ligand for voltage-sensitive sodium channels.
AuthorsBrown GB, Bradley RJ
JournalJ Neurosci Methods
PubMed ID2582213
Batrachotoxin and its derivatives have become important tools for the study of membrane excitability by virtue of their effects on voltage-sensitive sodium channels. Recent studies have shown that the 2,4-dimethylpyrrole carboxylate ester normally occurring in the 20-alpha position of batrachotoxin may be replaced by a benzoate moiety without loss of ... More
The effect of F-actin on the binding and hydrolysis of guanine nucleotide by Dictyostelium elongation factor 1A.
AuthorsEdmonds BT, Bell A, Wyckoff J, Condeelis J, Leyh TS
JournalJ Biol Chem
PubMed ID9553081
Indirect evidence implicates actin as a cofactor in eukaryotic protein synthesis. The present study directly examines the effects of F-actin on the biochemical properties of eukaryotic elongation factor 1A (eEF1A, formerly EF1alpha), a major actin-binding protein. The basal mechanism of eEF1A alone is determined under physiological conditions with the critical ... More
Fluorescent N-methylanthranilyl (Mantyl) tag for peptides: its application in subpicomole determination of kinins.
AuthorsAnumula KR, Schulz RP, Back N
JournalPeptides
PubMed ID1437709
Highly fluorescent N-methylanthranilyl (Mantyl) peptide derivatives were prepared by a one-step reaction with N-methylisatoic anhydride (MIA) for quantitative detection in HPLC. Reactions were carried out in an organic medium of acetonitrile-triethylamine, in aqueous alkaline sodium carbonate and sodium phosphate buffers. 4-Dimethylaminopyridine (DMAP) catalyzed specific mantylation of -NH2 groups of peptides ... More
Hydrolysis of GTP by p21NRAS, the NRAS protooncogene product, is accompanied by a conformational change in the wild-type protein: use of a single fluorescent probe at the catalytic site.
AuthorsNeal SE, Eccleston JF, Webb MR
JournalProc Natl Acad Sci U S A
PubMed ID2185475
2'(3')-O-(N-Methyl)anthraniloylguanosine 5'-triphosphate (mantGTP) is a fluorescent analogue of GTP that has similar properties to the physiological substrate in terms of its binding constant and the kinetics of its interactions with p21NRAS, the NRAS protooncogene product. There is a 3-fold increase in fluorescence intensity when mantGTP binds to p21NRAS. The rate ... More
Fluorescent analogs of UDP-glucose and their use in characterizing substrate binding by toxin A from Clostridium difficile.
AuthorsBhattacharyya S, Kerzmann A, Feig AL
JournalEur J Biochem
PubMed ID12135481
Uridine-5'-diphospho-1-alpha-d-glucose (UDP-Glc) is a common substrate used by glucosyltransferases, including certain bacterial toxins such as Toxins A and B from Clostridium difficile. Fluorescent analogs of UDP-Glc have been prepared for use in our studies of the clostridial toxins. These compounds are related to the methylanthraniloyl-ATP compounds commonly used to probe ... More
Rac "insert region" is a novel effector region that is implicated in the activation of NADPH oxidase, but not PAK65.
AuthorsFreeman JL, Abo A, Lambeth JD
JournalJ Biol Chem
PubMed ID8702687
The small GTPase Rac assembles with the cytosolic p47(phox) and p67(phox) and the membrane-associated flavocytochrome b558 to form the multicomponent respiratory burst oxidase. Mutation of amino acids in a region of Rac (residues 26-45), homologous to an effector region in Ras, was previously shown to interfere with Rac binding to ... More
Pathway of ATP hydrolysis by monomeric and dimeric kinesin.
AuthorsMoyer ML, Gilbert SP, Johnson KA
JournalBiochemistry
PubMed ID9454569
The ATPase mechanism for a monomeric Drosophila kinesin construct, K341, was determined by pre-steady-state kinetic methods and compared to dimeric kinesin, K401. We directly measured the kinetics of binding mantATP (a fluorescent ATP analog) to the microtubule K341 complex, the dissociation of K341 from the microtubule, and release of phosphate ... More
Investigation of the GTP-binding/GTPase cycle of Cdc42Hs using extrinsic reporter group fluorescence.
AuthorsNomanbhoy TK, Leonard DA, Manor D, Cerione RA
JournalBiochemistry
PubMed ID8605211
The overall goal of these studies was to examine the applicability of extrinsic reporter group fluorescence in monitoring the GTP-binding/GTPase cycle of a Ras-like GTP-binding protein. Toward this end, we have labeled the GTP-binding protein Cdc42Hs with the environmentally sensitive fluorophore succinimidyl 6-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoate (sNBD) at a single reactive lysine residue. ... More
Altering the reaction coordinate of the ATP sulfurylase-GTPase reaction.
AuthorsYang M, Leyh TS
JournalBiochemistry
PubMed ID9116005
ATP sulfurylase, isolated from Escherichia coli K-12, catalyzes and couples two reactions: the hydrolysis of GTP and the synthesis of APS (adenosine 5'-phosphosulfate). Its GTPase activity is regulated in response to ligand binding at the APS-forming active site. In particular, AMP mimics an intermediate-like form of the enzyme that increases ... More
Rac binding to p67(phox). Structural basis for interactions of the Rac1 effector region and insert region with components of the respiratory burst oxidase.
AuthorsNisimoto Y, Freeman JL, Motalebi SA, Hirshberg M, Lambeth JD
JournalJ Biol Chem
PubMed ID9228059
Activation of the respiratory burst oxidase involves the assembly of the membrane-associated flavocytochrome b558 with the cytosolic components p47(phox), p67(phox), and the small GTPase Rac. Herein, the interaction between Rac and p67(phox) is explored using functional and physical methods. Mutually facilitated binding (EC50) of Rac1 and p67(phox) within the NADPH ... More
Interaction of myosin subfragment 1 with fluorescent ribose-modified nucleotides. A comparison of vanadate trapping and SH1-SH2 cross-linking.
AuthorsCremo CR, Neuron JM, Yount RG
JournalBiochemistry
PubMed ID2110475
The environment near the ribose binding site of skeletal myosin subfragment 1 (S1) was investigated by use of two adenosine 5'-diphosphate analogues with fluorescent groups attached at the 2'- and 3'-hydroxyls of the ribose ring. We have compared steady-state and time-resolved fluorescent properties of the reversibly bound S1-nucleotide complexes and ... More
Alternating site mechanism of the kinesin ATPase.
AuthorsGilbert SP, Moyer ML, Johnson KA
JournalBiochemistry
PubMed ID9454568
The processivity of the microtubule-kinesin ATPase has been investigated using stopped-flow kinetic methods to measure the binding of each motor domain of the dimeric kinesin (K401) to the microtubule and the release of the fluorescent ADP analog, 2'(3')-O-(N-methylanthraniloyl)adenosine 5'-diphosphate (mantADP) from the active site of the motor domain. The results ... More
Partial G protein activation by fluorescent guanine nucleotide analogs. Evidence for a triphosphate-bound but inactive state.
AuthorsRemmers AE, Neubig RR
JournalJ Biol Chem
PubMed ID8617747
N-methyl-3'-O-anthranoyl (MANT) guanine nucleotide analogs are useful environmentally sensitive fluorescent probes for studying G protein mechanisms. Previously, we showed that MANT fluorescence intensity when bound to G protein was related to the degree of G protein activation where MANT-guanosine-5'-O-(3-thiotriphosphate) (mGTP gammaS) had the highest fluorescence followed by mGTP and mGDP, ... More