A recombinant polypeptide model of the second predicted nucleotide binding fold of the cystic fibrosis transmembrane conductance regulator is a GTP-binding protein.
AuthorsRandak C, Neth P, Auerswald EA, Assfalg-Machleidt I, Roscher AA, Hadorn HB, Machleidt W
JournalFEBS Lett
PubMed ID8946960
'Association reactions of a recombinant CFTR-NBF-2 polypeptide fused to glutathione S-transferase with guanine nucleotides were monitored quantitatively by recording the fluorescence enhancement of excited trinitrophenol (TNP)-labelled GTP after binding to NBF-2. Binding of TNP-GTP to the recombinant NBF-2 polypeptide was characterized by a Kd value of 3.9 microM. The corrected ... 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
Synthesis of 2',3'-O-(2,4,6-trinitrocyclohexadienylidine)guanosine 5'-triphosphate and a study of its inhibitory properties with adenylate cyclase.
AuthorsHoyer PB, Fletcher P, Haley BE
JournalArch Biochem Biophys
PubMed ID3954359
A fluorescent GTP analog 2',3'-O-(2,4,6-trinitrocyclohexadienylidine) guanosine 5'-triphosphate (TNP-GTP) has been prepared and some of its physical properties characterized. TNP-GTP was found to be a potent inhibitor of chick embryo heart adenylate cyclase as activated by guanyl 5'-(beta,gamma-imido)triphosphate (GppNHp), F-, and forskolin with Ki values in the 8-15 microM range. It ... More
Overexpression and purification of the carboxyl-terminal nucleotide-binding domain from mouse P-glycoprotein. Strategic location of a tryptophan residue.
AuthorsBaubichon-Cortay H, Baggetto LG, Dayan G, Di Pietro A
JournalJ Biol Chem
PubMed ID7916013
The cDNA encoding the C-terminal nucleotide-binding domain (NBD2) from mouse P-glycoprotein involved in multidrug resistance was obtained from adrenal cell mRNA and amplified by reverse transcriptase polymerase chain reaction. NBD2 was highly overexpressed in Escherichia coli in fusion with glutathione S-transferase and could be purified after efficient thrombin cleavage. Both ... More
GTP-induced membrane binding and ion channel activity of annexin VI: is annexin VI a GTP biosensor?
AuthorsKirilenko A, Golczak M, Pikula S, Buchet R, Bandorowicz-Pikula J
JournalBiophys J
PubMed ID11964259
Annexin VI (AnxVI) formed ion channels in planar lipid bilayers that were induced by the addition of millimolar guanosine 5'-triphosphate (GTP) at pH 7.4 and that were not accompanied by a penetration of the protein into the membrane hydrophobic region. GTP-influenced interactions of AnxVI with Ca2+/liposomes produced small structural alterations ... More
A chromophoric and fluorescent analog of GTP, 2',3'-O-(2,4,6-trinitrocyclohexadienylidene)-GTP, as a spectroscopic probe for the GTP inhibitory site of liver glutamate dehydrogenase.
AuthorsHiratsuka T
JournalJ Biol Chem
PubMed ID3988736
The ribose-modified chromophoric and fluorescent analog of ATP, 2',3'-O-(2,4,6-trinitrocyclohexadienylidene)-ATP (TNP-ATP) (Hiratsuka, T., and Uchida, K. (1973) Biochim. Biophys. Acta 320, 635-647 and Hiratsuka, T. (1976) Biochim. Biophys. Acta 453, 293-297) has been widely used as an ATP analog for various ATPases. Although the corresponding analog of GTP,2',3'-O-(2,4,6-trinitrocyclohexadienylidene)-GTP (TNP-GTP) should be ... More
Distances between the paclitaxel, colchicine, and exchangeable GTP binding sites on tubulin.
AuthorsHan Y, Malak H, Chaudhary AG, Chordia MD, Kingston DG, Bane S
JournalBiochemistry
PubMed ID9578547
Distances between the paclitaxel, colchicine, and exchangeable GTP binding sites on tubulin polymers have been probed using fluorescence spectroscopy. Techniques for measuring fluorescence resonance energy transfer (FRET) between fluorescent or chromophoric ligands for each binding site were employed. 2-Debenzoyl-2-(m-aminobenzoyl)paclitaxel (2-AB-PT) was the fluorophore ligand for the paclitaxel binding site; thiocolchicine, ... More
Comparative analysis of the ATP-binding sites of Hsp90 by nucleotide affinity cleavage: a distinct nucleotide specificity of the C-terminal ATP-binding site.
AuthorsSoti C, Vermes A, Haystead TA, Csermely P
JournalEur J Biochem
PubMed ID12755697
The 90-kDa heat shock protein (Hsp90) is a molecular chaperone that assists both in ATP-independent sequestration of damaged proteins, and in ATP-dependent folding of numerous targets, such as nuclear hormone receptors and protein kinases. Recent work from our lab and others has established the existence of a second, C-terminal nucleotide ... More