Bioluminescence resonance energy transfer from aequorin to a fluorophore: an artificial jellyfish for applications in multianalyte detection.
AuthorsDeo SK, Mirasoli M, Daunert S
JournalAnal Bioanal Chem
PubMed ID15731912
'In nature, the green light emission observed in the jellyfish Aequorea victoria is a result of a non-radiative energy transfer from the excited-state aequorin to the green fluorescent protein. In this work, we have modified the photoprotein aequorin by attaching selected fluorophores at a unique site on the protein. This ... More
Topology of the membrane domain of human erythrocyte anion exchange protein, AE1.
AuthorsFujinaga J, Tang XB, Casey JR
JournalJ Biol Chem
PubMed ID10037758
'Anion exchanger 1 (AE1) is the chloride/bicarbonate exchange protein of the erythrocyte membrane. By using a combination of introduced cysteine mutants and sulfhydryl-specific chemistry, we have mapped the topology of the human AE1 membrane domain. Twenty-seven single cysteines were introduced throughout the Leu708-Val911 region of human AE1, and these mutants ... More
Characterization of recombinant yeast dolichyl mannosyl phosphate synthase and site-directed mutagenesis of its cysteine residues.
AuthorsForsee WT, McPherson D, Schutzbach JS
JournalEur J Biochem
PubMed ID9108271
'Dolichyl mannosyl phosphate synthase is associated with membranes of the rough endoplasmic reticulum and catalyzes mannosyl transfer from GDP-mannose to the hydrophobic long-chain acceptor dolichyl-phosphate. The gene for the yeast enzyme encodes a protein with a molecular mass of 30.36 kDa containing three cysteine residues, at positions 93, 172 and ... More
Small vertical movement of a K+ channel voltage sensor measured with luminescence energy transfer.
AuthorsPosson DJ, Ge P, Miller C, Bezanilla F, Selvin PR
JournalNature
PubMed ID16094368
'Voltage-gated ion channels open and close in response to voltage changes across electrically excitable cell membranes. Voltage-gated potassium (Kv) channels are homotetramers with each subunit constructed from six transmembrane segments, S1-S6 (ref. 2). The voltage-sensing domain (segments S1-S4) contains charged arginine residues on S4 that move across the membrane electric ... More
Design consideration and probes for fluorescence resonance energy transfer studies.
AuthorsSinev M, Landsmann P, Sineva E, Ittah V, Haas E
JournalBioconjug Chem
PubMed ID10821651
'Spectroscopic properties of two newly synthesized water-soluble thiol-reactive fluorescent probes, 7-(iodoacetamido)-coumarin-4-carboxylic acid (I-Cca) and N-iodoacetyl-beta-(2-naphthyl)alanine (I-Nal), were characterized using single cysteine mutants of Escherichia coli adenylate kinase. Together with two known water-soluble thiol-reactive dyes (Lucifer yellow iodoacetamide and 5-iodoacetamidosalicylic acid) and as well, tryptophan residues (either native or inserted into ... More
Intracellular pH in renal tubules in situ: single-cell measurements by confocal laserscan microscopy.
AuthorsWeinlich M, Capasso G, Kinne RK
JournalPflugers Arch
PubMed ID8474854
'Confocal laserscan microscopy with a dual-excitation device was used to record intracellular pH (pHi) regulation in rat proximal convoluted tubules microperfused in vivo. Cells were loaded with the pH-sensitive, fluorescent dye 2'',7''-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Single cells could be distinguished within the tubules and separate measurements were possible. Application of an NH4Cl ... More
Topology of the region surrounding Glu681 of human AE1 protein, the erythrocyte anion exchanger.
AuthorsTang XB, Fujinaga J, Kopito R, Casey JR
JournalJ Biol Chem
PubMed ID9712881
'AE1 protein transports Cl- and HCO3- across the erythrocyte membrane by an electroneutral exchange mechanism. Glu681 of human AE1 may form part of the anion translocation apparatus and the permeability barrier. We have therefore studied the structure of the sequence surrounding Glu681, using scanning cysteine mutagenesis. Residues of the Ser643 ... More
Use of lucifer yellow iodoacetamide in a flow cytometric assay to measure cell surface free thiol.
AuthorsArcher JR, Badakere SS, Macey MG, Whelan MA
JournalBiochem Soc Trans
PubMed ID7758750
Sulfhydryl-selective fluorescence labeling of lipoprotein(a) reveals evidence for one single disulfide linkage between apoproteins(a) and B-100.
AuthorsSommer A, Gorges R, Kostner GM, Paltauf F, Hermetter A
JournalBiochemistry
PubMed ID1835655
Human lipoprotein(a) and low-density lipoprotein were labeled with two different sulfhydryl-selective fluorescence markers. The hydrophilic fluorophore lucifer yellow iodoacetamide and the apolar compound 6-acryloyl-2-(dimethylamino)naphthalene were used to derivatize free -SH groups in the lipoproteins. Three sulfhydryls could be detected in low-density lipoprotein, whereas only two cysteines were available in lipoprotein(a). ... More
A new potent HIV-1 reverse transcriptase inhibitor. A synthetic peptide derived from the interface subunit domains.
The biologically relevant and active forms of human immunodeficiency viruses type 1 and 2 reverse transcriptase found in infectious virions are heterodimers produced in a two-step dimerization process. Dimerization involves first the rapid association of the two subunits, followed by a slow conformational change yielding a fully active form. We ... More
Study of the conformation of DARPP-32, a dopamine- and cAMP-regulated phosphoprotein, by fluorescence spectroscopy.
DARPP-32 is a potent inhibitor of protein phosphatase 1 when it is phosphorylated on Thr34 by cAMP-dependent protein kinase. DARPP-32 is also phosphorylated on Ser45 and Ser102 by casein kinase II, resulting in a facilitation of phosphorylation by cAMP-dependent protein kinase. We have studied the conformation of recombinant rat DARPP-32 ... More
A novel potent strategy for gene delivery using a single peptide vector as a carrier.
AuthorsMorris MC, Chaloin L, Méry J, Heitz F, Divita G
JournalNucleic Acids Res
PubMed ID10446241
We have shown previously that a peptide, MPG, derived from the hydrophobic fusion peptide of HIV-1 gp41 and the hydrophilic nuclear localisation sequence of SV40 large T antigen, can be used as a powerful tool for the delivery of oligonucleotides into cultured cells. Now we extend the potential of MPG ... More
2,2'-Dithiobis(N-ethyl-spermine-5-carboxamide) is a high affinity, membrane-impermeant antagonist of the mammalian polyamine transport system.
AuthorsHuber M, Pelletier JG, Torossian K, Dionne P, Gamache I, Charest-Gaudreault R, Audette M, Poulin R
JournalJ Biol Chem
PubMed ID8910341
We have synthesized 2,2'-dithiobis(N-ethyl-spermine-5-carboxamide) (DESC), its thiol monomer (MESC), and the mixed MESC-cysteamine disulfide (DEASC) as potential inhibitors of polyamine transport in mammalian cells. DESC was the most potent antagonist of spermine transport in ZR-75-1 human breast cancer cells, with Ki values of 5. 0 +/- 0.7, 80 +/- 31, ... More
Subunit exchange of alphaA-crystallin.
AuthorsBova MP, Ding LL, Horwitz J, Fung BK
JournalJ Biol Chem
PubMed ID9368012
alpha-Crystallin, the major protein in the mammalian lens, is a molecular chaperone that can bind denaturing proteins and prevent their aggregation. Like other structurally related small heat shock proteins, each alpha-crystallin molecule is composed of an average of 40 subunits that can undergo extensive reorganization. In this study we used ... More