Modification of the actin interface of skeletal myosin subfragment-1 by treatment with dibromobimane.
AuthorsMornet D, Ue K, Chaussepied P, Morales MF
JournalEur J Biochem
PubMed ID3758077
'Recently, by treating the head portion of skeletal myosin subfragment-1 (S1) with the bifunctional agent dibromobimane, we introduced an intramolecular covalent cross-link which resulted in the stabilisation of an internal loop in the heavy chain structure of the head [Mornet et al. (1984) Proc. Natl Acad. Sci. USA 82, 1658-1662]. ... More
Biotinylated granulocyte/macrophage colony-stimulating factor analogues: effect of linkage chemistry on activity and binding.
'Biotinylated granulocyte/macrophage colony-stimulating factor (GM-CSF) analogues with different linkage chemistries and levels of conjugated biotin were synthesized by reacting recombinant human GM-CSF with sulfosuccinimidyl 6-biotinamidohexanoate or biotin hydrazide/1-[3-(dimethylamino)-propyl]-3-ethylcarbodiimide. These chemically reactive forms of biotin produced derivatives biotinylated at amine or carboxyl groups, respectively. Amine-derivatized analogues of 1.2 and 3.8 mol ... More
Zero-length crosslinking between subunits delta and I of the H(+)-translocating ATPase of chloroplasts.
AuthorsBeckers G, Berzborn RJ, Strotmann H
JournalBiochim Biophys Acta
PubMed ID1385976
'Treatment of spinach thylakoids with 1-ethyl-3-(dimethylaminopropyl)-carbodiimide (EDC)/N-hydroxysulfosuccinimide (sulfo-NHS) induced formation of a zero-length crosslink of an apparent molecular mass of 38 kDa. This product was shown, by immunodetection, to consist of subunit delta of CF1 and subunit I of CF0. The crosslink was isolated by preparative SDS gel electrophoresis and ... More
Carbodiimide as a tissue fixative in histamine immunohistochemistry and its application in developmental neurobiology.
AuthorsPanula P, Häppölä O, Airaksinen MS, Auvinen S, Virkamäki A
JournalJ Histochem Cytochem
PubMed ID3343510
'The object of this study was to develop an immunohistochemical method that could be used to study neuronal histamine, especially in nerve fibers and terminals where most previous methods have not been applicable. Three new antisera were produced in rabbits against conjugated histamine, and the fixative used in conjugation, 1-ethyl-3(3-diamethylaminopropyl)-carbodiimide ... More
Introduction of sulfhydryl groups into proteins at carboxyl sites.
AuthorsLin CM, Mihal KA, Krueger RJ
JournalBiochim Biophys Acta
PubMed ID2160279
'A two-step procedure for introduction of sulfhydryl groups at protein carboxyl groups is described. The resultant proteins contain 2-aminoethanethiol residues bound by amide linkages to the protein carboxyl groups. First an amide bond is formed between a carboxyl group of the protein and one of the amino groups of cystamine. ... More
Electroporation loading of calcium-sensitive dyes into the CNS.
AuthorsBonnot A, Mentis GZ, Skoch J, O'Donovan MJ
JournalJ Neurophysiol
PubMed ID15509647
'Calcium imaging of neural network function has been limited by the extent of tissue labeled or the time taken for labeling. We now describe the use of electroporation-an established technique for transfecting cells with genes-to load neurons with calcium-sensitive dyes in the isolated spinal cord of the neonatal mouse in ... More
Binding of muscimol-conjugated quantum dots to GABAC receptors.
AuthorsGussin HA, Tomlinson ID, Little DM, Warnement MR, Qian H, Rosenthal SJ, Pepperberg DR,
JournalJ Am Chem Soc
PubMed ID17147380
'Functionalization of highly fluorescent CdSe/ZnS core-shell nanocrystals (quantum dots, qdots) is an emerging technology for labeling cell surface proteins. We have synthesized a conjugate consisting of approximately 150-200 muscimols (a GABA receptor agonist) covalently joined to the qdot via a poly(ethylene glycol) (PEG) linker (approximately 78 ethylene glycol units) and ... More
Expressing functional domains of mouse calponin: involvement of the region around alanine 145 in the actomyosin ATPase inhibitory activity of calponin.
Authorsel-Mezgueldi M, Strasser P, Fattoum A, Gimona M
JournalBiochemistry
PubMed ID8619984
'Previously, we attributed the binding of F-actin to the 38-residue stretch of gizzard calponin encompassing the sequence A145-Y182 and postulated the hexapeptide motif VKYAEK, representing residues 142-147, as a putative actin-binding site [Mezgueldi, M., Fattoum, A., Derancourt, J. & Kassab, R. (1992) J. Biol. Chem. 267, 15943-15951]. Herein, the nature ... More
Prostate specific antigen biosensor based on long range surface plasmon-enhanced fluorescence spectroscopy and dextran hydrogel binding matrix.
AuthorsWang Y, Brunsen A, Jonas U, Dostálek J, Knoll W,
JournalAnal Chem
PubMed ID19894697
'A new biosensor based on surface plasmon-enhanced fluorescence spectroscopy (SPFS), which employs long-range surface plasmons (LRSP) and a photo-cross-linkable carboxymethyl dextran (PCDM) hydrogel binding matrix, is reported. LRSPs are surface plasmon modes that propagate along a thin metallic film with orders of magnitude lower damping compared to regular surface plasmons. ... More
Quantification of carboxyl groups in carbodiimide cross-linked collagen sponges.
AuthorsEveraerts F, Torrianni M, Hendriks M, Feijen J,
JournalJ Biomed Mater Res A
PubMed ID17595021
'Glutaraldehyde (GA) fixation of bioprosthetic tissue is a well adapted technique, with commercial products on the market for almost 40 years. Amine groups present in tissue react with GA to form different types of cross-links. An estimation of the degree of cross-linking of the tissue can be obtained by measuring ... More
A fluorescence resonance energy transfer-based approach for investigating late endosome-lysosome retrograde fusion events.
AuthorsKaufmann AM, Goldman SD, Krise JP,
JournalAnal Biochem
PubMed ID19109922
'Traditionally, lysosomes have been considered to be a terminal endocytic compartment. Recent studies suggest that lysosomes are quite dynamic, being able to fuse with other late endocytic compartments as well as with the plasma membrane. Here we describe a quantitative fluorescence energy transfer (FRET)-based method for assessing rates of retrograde ... More
Change in the actin-myosin subfragment 1 interaction during actin polymerization.
AuthorsChaussepied P, Kasprzak AA
JournalJ Biol Chem
PubMed ID2584238
'To better characterize the conformational differences of G- and F-actin, we have compared the interaction between G- and F-actin with myosin subfragment 1 (S1) which had part of its F-actin binding site (residues 633-642) blocked by a complementary peptide or "antipeptide" (Chaussepied, P., and Morales, M. F. (1988) Proc. Natl. ... More
Targeting glucose oxidase at aspartate and glutamate residues with organic two-electron redox mediators.
AuthorsBattaglini F, Koutroumanis M, English AM, Mikkelsen SR
JournalBioconjug Chem
PubMed ID7849073
'The bimolecular rate constants for the reactions of five organic two-electron redox mediators with reduced glucose oxidase (GOx) were determined by measuring voltammetric electrocatalytic currents at glassy carbon electrodes in the presence of excess glucose under anaerobic conditions. The mediators studied were thionine, brilliant cresyl blue, azure A, daunomycin, and ... More
Modulation of the immune response to pneumococcal type 14 capsular polysaccharide-protein conjugates by the adjuvant Quil A depends on the properties of the conjugates.
AuthorsVerheul AF, Versteeg AA, De Reuver MJ, Jansze M, Snippe H
JournalInfect Immun
PubMed ID2925240
'Streptococcus pneumoniae type 14 capsular polysaccharide-bovine serum albumin (S14PS-BSA) conjugates were prepared by water-soluble-carbodiimide-mediated condensation with or without the use of N-hydroxy-sulfosuccinimide. The immunogenicities of the capsular polysaccharide (S14PS) and of the conjugates were studied in (CBA/N x BALB/c)F1 mice and in female BALB/c mice. The response in these mice ... More
Binding of heavy-chain and essential light-chain 1 of S1 to actin depends on the degree of saturation of F-actin filaments with S1.
AuthorsAndreev OA, Borejdo J
JournalBiochemistry
PubMed ID7578092
'The interaction of heavy-chain isoforms of myosin subfragment-1 with actin was examined by cross-linking with carbodiimide (EDC). The heavy chain of S1 could be cross-linked to a single actin molecule through sites on either 20 or 50 kDa proteolytic domains, resulting in complexes which migrated in an 8% polyacrylamide gel ... More
Fast electron transfer from cytochrome c6 and plastocyanin to photosystem I of Chlamydomonas reinhardtii requires PsaF.
AuthorsHippler M, Drepper F, Farah J, Rochaix JD
JournalBiochemistry
PubMed ID9174349
'To study the function of the PsaF subunit of photosystem I (PSI), the interactions between plastocyanin or cytochrome c6 and PSI isolated from wild-type and a PsaF-deficient mutant of Chlamydomonas reinhardtii have been examined using cross-linking techniques and flash absorption spectroscopy. We show that efficient electron transfer from both plastocyanin ... More
Actin assembly induced by polylysine beads or purified phagosomes: quantitation by a new flow cytometry assay.
AuthorsDefacque H, Egeberg M, Antzberger A, Ansorge W, Way M, Griffiths G
JournalCytometry
PubMed ID10942896
'BACKGROUND: Actin assembly on biological membranes is a poorly understood process. We have previously shown that phagosomal membranes could induce actin assembly in the presence of thymosin beta4 (an actin sequestering protein that inhibits nonspecific nucleation), via the barbed ends of actin filaments. METHODS: Here, we have developed an in ... More
Carbodiimide modification enhances activity of pig pancreatic phospholipase A2.
AuthorsFerreira JP, Sasisekharan R, Louie O, Langer R
JournalEur J Biochem
PubMed ID8055932
'Pig phospholipase A2, pig iso-phospholipase A2 and bovine pancreatic phospholipase A2 were reacted in solution with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, in the presence of N-hydroxysulfosuccinimide, at pH 7. The influence of micellar protectants was analyzed. In the presence of n-hexadecylphosphocholine, the losses of activity in micellar diheptanoyl-lecithin were 80, 35, and 10% in ... More
Molecular contacts between nebulin and actin: cross-linking of nebulin modules to the N-terminus of actin.
AuthorsShih CL, Chen MJ, Linse K, Wang K
JournalBiochemistry
PubMed ID9048566
'Nebulin, a giant actin binding protein, coextends with actin and is thought to form a composite thin filament in the skeletal muscle sarcomere. To understand the molecular interactions between nebulin and actin, we have applied chemical cross-linking techniques to define molecular contacts between actin and ND8, a two-module nebulin fragment ... More
Spatial organization of template polynucleotides on the ribosome determined by fluorescence methods.
AuthorsBakin AV, Borisova OF, Shatsky IN, Bogdanov AA
JournalJ Mol Biol
PubMed ID1717698
'The spatial organization of template polynucleotides on the ribosome and the dynamics of their interaction with 30 S subunits have been studied by fluorescence spectroscopy. The topography of the mRNA in the ribosome has been determined using singlet-singlet energy transfer. This method has allowed us to estimate distances between donors ... More
Amide cross-linking: an alternative to glutaraldehyde fixation.
AuthorsGirardot JM, Girardot MN
JournalJ Heart Valve Dis
PubMed ID8894992
'BACKGROUND AND AIMS OF THE STUDY: A new fixation method for bioprosthetic tissues is being developed, which does not utilize the standard glutaraldehyde treatment. This method, referred to as Ultifix, uses a coupler and a coupling enhancer with or without one or more coupling agents. It fixes the tissue by ... More
Zero-length crosslinking procedure with the use of active esters.
AuthorsGrabarek Z, Gergely J
JournalAnal Biochem
PubMed ID2344038
'A two-step zero-length crosslinking procedure for studying protein-protein complexes has been developed. One component of a complex is briefly incubated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) in the presence of N-hydroxysuccinimide resulting in the conversion of some of the protein carboxyls into succinimidyl esters. The reaction is stopped by addition of beta-mercaptoethanol and ... More
Oxidation-triggered release of fluorescent molecules or drugs from mesoporous Si microparticles.
AuthorsWu EC, Park JH, Park J, Segal E, Cunin F, Sailor MJ,
JournalACS Nano
PubMed ID19206408
'The fluorescent dye Alexa Fluor 488 or the anticancer drug doxorubicin is attached to the surface and inner pore walls of mesoporous Si particles by covalent attachment, and the oxidation-induced release of each molecule is studied. The molecules are bound to the Si matrix using a 10-undecenoic acid linker, which ... More
Ni(II) and Ni(I) forms of pentaalkylamide derivatives of cofactor F430 of Methanobacterium thermoautotrophicum.
AuthorsHamilton CL, Ma L, Renner MW, Scott RA
JournalBiochim Biophys Acta
PubMed ID1648401
'A series of pentaalkylamide forms of F430 and of its 12,13-diepimer have been generated and characterized. Carbodiimide-assisted N-hydroxysulfosuccinimide activation of all five peripheral carboxylates of the F430 macrocycle allows nucleophilic attack by a number of primary amines (RNH2, R- = CH3-, CH3CH2-, CF3CH2-, CH3(CH2)3-) generating the pentaalkylamide derivatives. The identity ... More
Succinylated polylysine as a possible link between an antibody molecule and deferoxamine.
'Modification of antibodies with chelating polymers may be helpful for radioimmunoimaging, radioimmunotherapy, and NMR tomography. Succinylated polylysine was activated with carbodiimide/N-hydroxysulfosuccinimide in dimethyl sulfoxide and isolated as a dry solid. Sulfosuccinimide-esterified polymer was used for the two-stage coupling of an amino-containing chelating agent (deferoxamine) to monoclonal R11D10 (IgG) or its ... More
Protein-heparin interactions measured by BIAcore 2000 are affected by the method of heparin immobilization.
AuthorsOsmond RI, Kett WC, Skett SE, Coombe DR
JournalAnal Biochem
PubMed ID12423639
'Surface plasmon resonance (SPR) biosensors such as the BIAcore 2000 are a useful tool for the analysis of protein-heparin interactions. Generally, biotinylated heparin is captured on a streptavidin-coated surface to create heparinized surfaces for subsequent binding analyses. In this study we investigated three commonly used techniques for the biotinylation of ... More
Stability of water-soluble carbodiimides in aqueous solution.
AuthorsGilles MA, Hudson AQ, Borders CL
JournalAnal Biochem
PubMed ID2158246
'A dimethylbarbituric acid reagent has been used to follow the kinetics of loss of two water-soluble carbodiimides, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and the structurally related 1-ethyl-3-(4-azonia-4,4-dimethylpentyl) carbodiimide (EAC), in aqueous solution as a function of pH and added chemical reagents. In 50 mM 2-(N-morpholino)ethanesulfonic acid at 25 degrees C, EDC has ... More
Water-soluble carbodiimide for the fluorescent measurement of the carboxyl group produced by enzyme reactions.
AuthorsKobayashi M, Chiba Y
JournalAnal Biochem
PubMed ID8080075
'Fluorescent measurement of the carboxyl group was achieved using water-soluble carbodiimide, EDC (1-ethyl-3-(3- dimethylaminopropyl)carbodiimide). Detection of the active intermediate from the glucuronic acid-EDC reaction mixture using the OPA (o-phthalaldehyde) reagent described in a previous paper was greatly improved to about 4 ng/tube (17 pmol) glucuronic acid owing to a modification ... More
The functional role of carboxyl residues in an acyl-CoA dehydrogenase.
AuthorsFrerman FE, Mielke D, Huhta K
JournalJ Biol Chem
PubMed ID7354089
Reaction of insulin with ethyl glycinate and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide.
AuthorsOzawa H
JournalBiochemistry
PubMed ID5442178
Altered arrangement of the DNA in injection-defective lambda bacteriophage.
AuthorsThomas JO, Sternberg N, Weisberg R
JournalJ Mol Biol
PubMed ID355644
The use of carbodiimides in the preparation of immunizing conjugates.
AuthorsBauminger S, Wilchek M
JournalMethods Enzymol
PubMed ID6999295
In situ analysis of carboxyl and sulfhydryl groups of extracellular polymeric secretions by confocal laser scanning microscopy.
AuthorsKawaguchi T, Decho AW
JournalAnal Biochem
PubMed ID12009705
Constraints on the flexibility of bacteriorhodopsin's carboxyl-terminal tail at the purple membrane surface.
AuthorsRenthal R, Dawson N, Tuley J, Horowitz P
JournalBiochemistry
PubMed ID6830763
Copper-free click chemistry for highly luminescent quantum dot conjugates: application to in vivo metabolic imaging.
AuthorsBernardin A, Cazet A, Guyon L, Delannoy P, Vinet F, Bonnaffé D, Texier I,
JournalBioconjug Chem
PubMed ID20222737
Quantum dots (QD) are inorganic nanocrystals with outstanding optical properties, specially suited for biological imaging applications. Their attachment to biomolecules in mild aqueous conditions for the design of bioconjugates is therefore highly desirable. 1,3-dipolar [3 + 2] cycloaddition between azides and terminal alkynes ( ... More
Enhancement by N-hydroxysulfosuccinimide of water-soluble carbodiimide-mediated coupling reactions.
AuthorsStaros JV, Wright RW, Swingle DM
JournalAnal Biochem
PubMed ID3740412
Water-soluble carbodiimides are frequently employed in coupling or conjugation reactions, e.g., to link a peptide immunogen to a carrier protein. However, their utility is limited by low coupling yields obtained under some conditions. We have found that addition of N-hydroxysulfosuccinimide to such reactions can greatly enhance the yields obtained. ... More
An active-site carboxyl group in liquefying alpha-amylase: specific chemical modification.
AuthorsKochhar S, Dua RD
JournalBiosci Rep
PubMed ID6332649
Bacillus amyloliquefaciens alpha-amylase activity is pH-dependent and the plot log (Vmax/Km) versus pH implicated a carboxyl group of aspartic acid/glutamic acid at the active site. Chemical modification of alpha-amylase with EDC confirmed this view. Further, analysis of inactivation kinetics showed that modification of a single carboxyl group led to complete ... More
Labeling of free carboxyl groups.
AuthorsYasaka Y, Tanaka M
JournalJ Chromatogr B Biomed Appl
PubMed ID7820274
The latest trends in the labeling of free carboxyl groups for high-performance liquid chromatography are reviewed. The labeling reagents for fluorescence detection are mainly discussed according to their reaction type (or functional group). Attention is also paid to the reagents used for ultraviolet detection and for enantiomeric separation. The reactivity ... More
A cleavable cross-linking reaction for protein carboxyl groups.
AuthorsRenthal R
JournalInt J Pept Protein Res
PubMed ID6311762
L-(+)-tartaric acid dihydrazide was coupled to the purple membrane from Halobacterium halobium with a water soluble carbodiimide. Gel electrophoresis analysis of the product revealed the formation of bacteriorhodopsin dimers, trimers and higher polymers. Most of the cross-links were removed by treatment with papain, demonstrating involvement of the carboxyl-rich carboxyl-terminal region ... More
Preparation of novel cyclosporin A derivatives.
AuthorsPaprica PA, Margaritis A, Petersen NO
JournalBioconjug Chem
PubMed ID1616947
The hydroxyl group on the 2-N-methyl-(R)-((E)-2-butenyl)-4-methyl-L-threonine residue of cyclosporin A was protected by acetylation, then the double bond on the same amino acid residue was oxidatively cleaved using a periodate/permanganate reagent. The resultant derivative of cyclosporin A contained a carboxylic acid group which was subsequently reacted with the nucleophiles 5-(aminoacetamido)fluorescein ... More
Methods for determining the modification of protein thiols by reactive lipids.
AuthorsOh J, Johnson MS, Landar A,
JournalMethods Cell Biol
PubMed ID17445707
Biotin-based tagging techniques have been applied with success to monitor the posttranslational modification of reactive protein thiols in cell signaling proteins and enzymes by ROS/RNS and electrophilic compounds. One of the most versatile techniques is to attach biotin covalently to thiol-reactive molecules, and label thiol-containing proteins. After separation ... More
Inactivation of Ca2(+)-, Na+K(+)-, and H+K(+)-ATPases with a carbodiimide derivative of ATP.
AuthorsMurphy AJ
JournalFEBS Lett
PubMed ID2158904
The gamma-P adduct of ATP with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (ATP-EDC) was synthesized and incubated with the Ca-ATPase of sarcoplasmic reticulum with the result that time-dependent complete loss of the enzyme's activity occurred. The inactivation required calcium and magnesium while ATP had a protective effect. ATP-EDC incubation with the NaK-ATPase and HK-ATPase produced ... More
Use of water-soluble 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide for the fluorescent determination of uronic acids and carboxylic acids.
AuthorsKobayashi M, Ichishima E
JournalAnal Biochem
PubMed ID2278381
Reaction between glucuronic acid and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was monitored by the o-phthalaldehyde (OPA) method, which was developed for the fluorescent assay of compounds containing an amino group. About 1 nmol of glucuronic acid was detected by this method. This EDC-OPA method was effective in detecting not only acidic sugar but ... More
Biotin-protein bond: instability and structural modification to provide stability for in vivo applications.
AuthorsMock DM, Bogusiewicz A,
JournalMethods Mol Biol
PubMed ID18287661
Biotinylation of proteins is a powerful tool for investigating biological phenomenon, both in vitro and in vivo. Biotinylating reagents that form covalent bonds with several types of amino acid residues are commercially available. However, most, if not all, of these commercially available biotinylating agents produce biotin-protein bonds that are susceptible ... More
Shape effects of nanoparticles conjugated with cell-penetrating peptides (HIV Tat PTD) on CHO cell uptake.
AuthorsZhang K, Fang H, Chen Z, Taylor JS, Wooley KL,
JournalBioconjug Chem
PubMed ID18690739
In order to probe the nanoparticle shape/size effect on cellular uptake, a spherical and two cylindrical nanoparticles, whose lengths were distinctively varied, were constructed by the selective cross-linking of amphiphilic block copolymer micelles. Herein, we demonstrate that, when the nanoparticles were functionalized with the protein transduction domain of human immunodeficiency ... More
A model system for convenient fluorescent labeling of sugar chain in taka-amylase A.
AuthorsKobayashi M, Sasaki Y, Chiba Y
JournalBiosci Biotechnol Biochem
PubMed ID9404062
A convenient detection of sugar chains in Taka-amylase A (TAA) was done by using 40 micrograms of enzyme, where a decrease in the UV absorption of NaIO4 during the periodate oxidation reaction was monitored. The periodate-oxidized sugar chain was labeled with a fluorescent reagent, N-1-ethylenediaminonaphthalene (EDAN), by incubation at pH ... More
A novel FRET approach for in situ investigation of cellulase-cellulose interaction.
AuthorsWang L, Wang Y, Ragauskas AJ,
JournalAnal Bioanal Chem
PubMed ID20694721
A novel real-time in situ detection method for the investigation of cellulase-cellulose interactions based on fluorescence resonance energy transfer (FRET) has been developed. FRET has been widely used in biological and biophysical fields for studies related to proteins, nucleic acids, and small biological molecules. Here, we report the efficient labeling ... More
An actin-binding site on the 20K fragment of myosin subfragment 1.
AuthorsSutoh K
JournalBiochemistry
PubMed ID7138830
Myosin subfragment 1 (S1) was covalently labeled with a fluorescent dye, N-[7-(dimethylamino)-4-methyl-3-coumarinyl]maleimide (DACM), and then digested by trypsin to cleave S1 heavy chain into fragments. The DACM-labeled and trypsin-treated S1 was complexed with F-actin and treated with a zero-length cross-linker, 1-ethyl-3[3-(dimethylamino)propyl] carbodiimide (EDC). The cross-linking reaction generated a covalently linked ... More
GABAC receptor binding of quantum-dot conjugates of variable ligand valency.
AuthorsGussin HA, Tomlinson ID, Muni NJ, Little DM, Qian H, Rosenthal SJ, Pepperberg DR,
JournalBioconjug Chem
PubMed ID20715850
Highly fluorescent CdSe quantum dots (qdots) can serve as a platform for tethering multiple copies of a receptor-targeted ligand, affording study of how the level of multivalency affects receptor binding. We previously showed that qdots conjugated with long PEG chains terminated by muscimol, a known GABA(C) agonist, exhibit specific binding ... More
Single cell kinetics of intracellular, nonviral, nucleic acid delivery vehicle acidification and trafficking.
AuthorsKulkarni RP, Mishra S, Fraser SE, Davis ME,
JournalBioconjug Chem
PubMed ID16029041
Mechanistic understanding of the intracellular trafficking of nonviral nucleic acid delivery vehicles remains elusive. A live, single cell-based assay is described here that is used to investigate and quantitate the spatiotemporal, intracellular pH microenvironment of polymeric-based nucleic acid delivery vehicles. Polycations such as polyethylenimine (PEI), poly-l-lysine (PLL), beta-cyclodextrin-containing polymers lacking ... More
SOI optical microring resonator with poly(ethylene glycol) polymer brush for label-free biosensor applications.
AuthorsDe Vos K, Girones J, Popelka S, Schacht E, Baets R, Bienstman P,
JournalBiosens Bioelectron
PubMed ID19200711
Label-free monitoring of biomolecular interactions has become of key importance for the emerging proteomics field. Monitoring real time interaction kinetics and high throughput screening of complex samples is of major importance for a variety of applications. We previously reported the use of Silicon-on-Insulator photonics microring resonators for cheap disposable biosensors ... More
Nanowire-transistor based ultra-sensitive DNA methylation detection.
Accurate detection of DNA methylation at specific gene transcription sites is important to identify potential tumor formation since this epigenetic alteration may result in silencing of tumor suppressor genes that protect against tumor formation or that repair damaged DNA. Current technologies used in DNA methylation detection are complicated and time ... More
Clicked bivalent ligands containing curcumin and cholesterol as multifunctional abeta oligomerization inhibitors: design, synthesis, and biological characterization.
In our effort to develop multifunctional compounds that cotarget beta-amyloid oligomers (AbetaOs), cell membrane/lipid rafts (CM/LR), and oxidative stress, a series of bivalent multifunctional Abeta oligomerization inhibitors (BMAOIs) containing cholesterol and curcumin were designed, synthesized, and biologically characterized as potential treatments for Alzheimer's disease (AD). The in vitro assay results ... More
Mapping of the protein import machinery in the mitochondrial outer membrane by crosslinking of translocation intermediates.
AuthorsSöllner T, Rassow J, Wiedmann M, Schlossmann J, Keil P, Neupert W, Pfanner N
JournalNature
PubMed ID1530986
Mitochondria contain a complex machinery for the import of nuclear-encoded proteins. Receptor proteins exposed on the outer membrane surface are required for the specific binding of precursor proteins to mitochondria, either by binding of cytosolic signal recognition factors or by direct recognition of the precursor polypeptides. Subsequently, the precursors are ... More
Stabilization of human prostatic acid phosphatase by coupling with chondroitin sulfate.
AuthorsLuchter-Wasylewska E, Dulinska J, Ostrowski WS, Torchilin VP, Trubetskoy VS
JournalBiotechnol Appl Biochem
PubMed ID1905136
Human prostatic acid phosphatase (PAP) (EC 3.1.3.2) was covalently linked to chondroitin sulfate A from whale cartilage. In order to bind the protein amino groups with the preactivated carboxyl groups of chondroitin sulfate, 1-ethyl-3-(3'-dimethylaminepropyl)carbodiimide and N-hydroxysulfosuccinimide were used as coupling agents. The product was soluble and enzymatically active. The activity ... More
Identification of the amino acids involved in the functional interaction between photosystem I and ferredoxin from Synechocystis sp. PCC 6803 by chemical cross-linking.
AuthorsLelong C, Sétif P, Lagoutte B, Bottin H
JournalJ Biol Chem
PubMed ID8144501
Ferredoxin isolated from the cyanobacterium Synechocystis sp. PCC 6803 has been chemically cross-linked to purified photosystem I from the same organism. The reaction was catalyzed by N-ethyl-3-(3-dimethylaminopropyl)carbodiimide in the presence of N-hydroxysulfosuccinimide. A short reaction time and neutral pH values can be used in the presence of the two reagents, ... More
Bioconjugated quantum dots for multiplexed and quantitative immunohistochemistry.
AuthorsXing Y, Chaudry Q, Shen C, Kong KY, Zhau HE, Chung LW, Petros JA, O'Regan RM, Yezhelyev MV, Simons JW, Wang MD, Nie S
JournalNat Protoc
PubMed ID17546006
Bioconjugated quantum dots (QDs) provide a new class of biological labels for evaluating biomolecular signatures (biomarkers) on intact cells and tissue specimens. In particular, the use of multicolor QD probes in immunohistochemistry is considered one of the most important and clinically relevant applications. At present, however, clinical applications of QD-based ... More
Au nanoparticle conjugation for impedance and capacitance signal amplification in biosensors.
AuthorsWang J, Profitt JA, Pugia MJ, Suni II
JournalAnal Chem
PubMed ID16536410
Amplification of the electrochemical impedance and capacitance signals in a biosensor is demonstrated for the model fluorescein/anti-fluorescein system. Following immobilization of fluorescein onto Au through formation of a self-assembled monolayer, goat anti-fluorescein conjugated with 10-nm Au nanoparticles is introduced into the system. This results in an increase in the capacitance ... More
Dual-colour microscopy of single fluorophores bound to myosin interacting with fluorescently labelled actin using anti-Stokes fluorescence.
AuthorsSaito K, Tokunaga M, Iwane AH, Yanagida T
JournalJ Microsc
PubMed ID9450329
We have refined prismless total internal reflection fluorescence microscopy with extremely low background to visualize single fluorophores attached to protein molecules interacting with a filamentous biopolymer labelled with different colour fluorophores. By using Stokes and anti-Stokes fluorescence, two different colour fluorescences from two different colour fluorophores excited with a single ... More
A hyaluronic acid-taxol antitumor bioconjugate targeted to cancer cells.
AuthorsLuo Y, Ziebell MR, Prestwich GD
JournalBiomacromolecules
PubMed ID11710102
A cell-targeted polymeric prodrug prepared from Taxol and chemically modified hyaluronic acid (HA) was evaluated in vitro. Herein we report four results in support of the selective uptake and targeted toxicity of the HA-Taxol prodrug. First, a fluorescently labeled HA-Taxol (FITC-HA-Taxol) was synthesized and used to demonstrate cell-specific binding and ... More
Assembly of the preprotein receptor MOM72/MAS70 into the protein import complex of the outer membrane of mitochondria.
AuthorsSchlossmann J, Neupert W
JournalJ Biol Chem
PubMed ID7592965
Membrane integration and assembly of MOM72 from Neurospora crassa and its yeast homolog MAS70 was studied with isolated mitochondria. After synthesis in vitro, the precursors of MOM72/MAS70 are tightly folded and expose only their N-terminal amino acid residues comprising the targeting and the membrane anchor domain. Insertion of the protein ... More
Interaction of myosin with F-actin: time-dependent changes at the interface are not slow.
AuthorsVan Dijk J, Céline F, Barman T, Chaussepied P
JournalBiophys J
PubMed ID10827986
The kinetics of formation of the actin-myosin complex have been reinvestigated on the minute and second time scales in sedimentation and chemical cross-linking experiments. With the sedimentation method, we found that the binding of the skeletal muscle myosin motor domain (S1) to actin filament always saturates at one S1 bound ... More
A single myosin head can be cross-linked to the N termini of two adjacent actin monomers.
AuthorsBonafé N, Chaussepied P
JournalBiophys J
PubMed ID7787098
Myosin subfragment-1 (S1) can be cross-linked to two actin monomers by 1-ethyl-3-[3-(dimethylamino)-propyl]-carbodiimide only when F-actin is in excess over S1. Electron micrographs of the covalent actin2-S1 complex showed that S1 was cross-linked to two adjacent monomers of the same actin filament. Cross-linking experiments with pre-proteolyzed S1 derivatives in combination with ... More
Modification of essential carboxyl group in rabbit muscle phosphorylase by water-soluble carbodiimide.
AuthorsTakagi S, Kobayashi M, Matsuda K
JournalJ Biochem (Tokyo)
PubMed ID2504706
Water-soluble carbodiimide (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide) (EDC) and glycine ethyl ester (GEE) as a nucleophile were used to modify the essential carboxyl group of phosphorylases. The inactive b form of the muscle phosphorylase was modified faster than the active a form and potato phosphorylases. Use of N,N,N',N'-tetramethyl-ethylenediamine (TEMED)-HCl buffer system (pH 6.2) resulted ... More
Fluorescent labeling of bacteriorhodopsin: implications for helix connections.
AuthorsRenthal R, Cothran M, Dawson N, Harris GJ
JournalBiochim Biophys Acta
PubMed ID3101736
Purple membrane from Halobacterium halobium was reacted with dansyl (5-dimethylamino-1-naphthalenyl fluorescent labels that have specificity for different protein side chains of bacteriorhodopsin. Dansyl chloride was found to react primarily with Lys-41. Dansyl hydrazine was coupled, with water-soluble carbodiimide, to Glu-74 and/or Asp-85, which was the major modified site after papain-cleavage ... More
Affinity cytochemical labeling of glomerular basement membrane anionic sites using specific biotinylation and colloidal gold probes.
AuthorsBertolatus JA
JournalJ Histochem Cytochem
PubMed ID2303703
To label specific anionic sites on glomerular capillary wall structures, biotin was covalently linked to sialic acid residues by sequential treatment with mild peroxidation and biotin hydrazide, while carboxyl groups were biotinylated by exposure to the combination of biotin hydrazide and a water-soluble carbodiimide reagent. Optimal specific labeling of rat ... More
5-Lipoxygenase interacts with coactosin-like protein.
We have recently identified coactosin-like protein (CLP) in a yeast two-hybrid screen using 5-lipoxygenase (5LO) as a bait. In this report, we demonstrate a direct interaction between 5LO and CLP. 5LO associated with CLP, which was expressed as a glutathione S-transferase fusion protein, in a dose-dependent manner. Coimmunoprecipitation experiments using ... More
Solution structure of the epsilon subunit of the F1-ATPase from Escherichia coli and interactions of this subunit with beta subunits in the complex.
AuthorsWilkens S, Capaldi RA
JournalJ Biol Chem
PubMed ID9756905
The solution structure of the epsilon subunit of the Escherichia coli F1-ATPase has been determined by NMR spectroscopy. This subunit has a two-domain structure with an N-terminal 10-stranded beta sandwich and a C-terminal antiparallel two alpha-helix hairpin, as described previously (Wilkens, S., Dahlquist, F. W., McIntosh, L. P., Donaldson, L. ... More
Two Caenorhabditis elegans actin depolymerizing factor/cofilin proteins, encoded by the unc-60 gene, differentially regulate actin filament dynamics.
AuthorsOno S, Benian GM
JournalJ Biol Chem
PubMed ID9452511
The Caenorhabditis elegans unc-60 gene encodes two actin depolymerizing factor/cofilin proteins which are implicated in the regulation of actin filament assembly in body wall muscle. We examined the interaction of recombinant UNC-60A and B proteins with actin and found that they differentially regulate actin filament dynamics. Co-pelleting assays with F-actin ... More
Thiol modification as a probe of conformational forms of the F1 ATPase of Escherichia coli and of the structural asymmetry of its beta subunits.
AuthorsStan-Lotter H, Bragg PD
JournalEur J Biochem
PubMed ID2867900
The sulfhydryl groups of soluble and membrane-bound F1 adenosine triphosphatase of Escherichia coli were modified by reaction with the fluorescent thiol reagents 5-iodoacetamidofluorescein, 2-[(4'-iodoacetamido)anilino]naphthalene-6-sulfonic acid 4-[N-(iodoacetoxy)ethyl-N-methyl]amino-7-nitrobenzo-2-oxa-1,3-d iaz ole and 2-[(4'-maleimidyl)anilino]naphthalene-6-sulfonic acid. Whereas gamma and delta subunits were always labeled by these reagents, the beta subunit reacted preferentially in the soluble ... More
Membrane topography and near-neighbor relationships of the mitochondrial ATP synthase subunits e, f, and g.
AuthorsBelogrudov GI, Tomich JM, Hatefi Y
JournalJ Biol Chem
PubMed ID8702768
The well characterized subunits of the bovine ATP synthase complex are the alpha, beta, gamma, delta, and epsilon subunits of the catalytic sector, F1; the ATPase inhibitor protein; and subunits a, b, c, and d, OSCP (oligomycin sensitivity-conferring protein), F6, and A6L, which are present in the membrane sector, F0, ... More
Detection of fluorescently labeled actin-bound cross-bridges in actively contracting myofibrils.
AuthorsCooper WC, Chrin LR, Berger CL
JournalBiophys J
PubMed ID10692330
Myosin subfragment 1 (S1) can be specifically modified at Lys-553 with the fluorescent probe FHS (6-[fluorescein-5(and 6)-carboxamido]hexanoic acid succinimidyl ester) (Bertrand, R., J. Derancourt, and R. Kassab. 1995. Biochemistry. 34:9500-9507), and solvent quenching of FHS-S1 with iodide has been shown to be sensitive to actin binding at low ionic strength ... More
Tbeta 4 is not a simple G-actin sequestering protein and interacts with F-actin at high concentration.
AuthorsCarlier MF, Didry D, Erk I, Lepault J, Van Troys ML, Vandekerckhove J, Perelroizen I, Yin H, Doi Y, Pantaloni D
JournalJ Biol Chem
PubMed ID8621582
Thymosin beta 4 is acknowledged as a major G-actin binding protein maintaining a pool of unassembled actin in motile vertebrate cells. We have examined the function of Tbeta 4 in actin assembly in the high range of concentrations (up to 300 micron) at which Tbeta 4 is found in highly ... More
Carbohydrate-carbohydrate binding of ganglioside to integrin alpha(5) modulates alpha(5)beta(1) function.
AuthorsWang X, Sun P, Al-Qamari A, Tai T, Kawashima I, Paller AS
JournalJ Biol Chem
PubMed ID11118433
Gangliosides GT1b and GD3, components of keratinocyte membranes, inhibit keratinocyte adhesion to fibronectin. Although ganglioside sialylation is known to be important, the mechanism of inhibition is unknown. Using purified insect recombinant alpha(5) and beta(1) proteins and alpha(5)beta(1) integrin from lysed keratinocyte-derived SCC12 cells, we have shown that GT1b and GD3 ... More
Confocal microscopic analysis of the interaction between cisplatin and the copper transporter ATP7B in human ovarian carcinoma cells.
AuthorsKatano K, Safaei R, Samimi G, Holzer A, Tomioka M, Goodman M, Howell SB
JournalClin Cancer Res
PubMed ID15240550
Some cisplatin (DDP)-resistant cells overexpress the copper export transporter ATP7B, and cells molecularly engineered to overexpress ATP7B are resistant to DDP. The interaction of Cu with ATP7B normally triggers its relocalization from the perinuclear region to more peripheral vesicles. To investigate the interaction of DDP with ATP7B, we examined the ... More
Specific carbodiimide-binding mechanism for the selective modification of the aspartic acid-101 residue of lysozyme in the carbodiimide-amine reaction.
AuthorsKuroki R, Yamada H, Imoto T
JournalJ Biochem (Tokyo)
PubMed ID3711072
A mechanism for the selective modification of Asp-101 in hen egg-white lysozyme with an amine nucleophile catalyzed by 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDC) was investigated using ethanolamine as a nucleophile at pH 5.0 and room temperature. In the presence of N-acetyl-D-glucosamine (NAG) and its oligomers [(NAG)n, n = 2 and 3] under ... More
Functional significance of the binding of one myosin head to two actin monomers.
AuthorsLheureux K, Chaussepied P
JournalBiochemistry
PubMed ID7547872
The functional significance of the interaction of one myosin head (S1) with two actin monomers was investigated by comparing the properties of the cross-linked monomeric and filamentous actin-S1 complexes. S1 was cross-linked to monomeric actin (G-actin) either in the absence or in the presence of DNase I by 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide. The ... More
Direct measurements of multiple adhesive alignments and unbinding trajectories between cadherin extracellular domains.
AuthorsSivasankar S, Gumbiner B, Leckband D
JournalBiophys J
PubMed ID11259289
Direct measurements of the interactions between antiparallel, oriented monolayers of the complete extracellular region of C-cadherin demonstrate that, rather than binding in a single unique orientation, the cadherins adhere in three distinct alignments. The strongest adhesion is observed when the opposing extracellular fragments are completely interdigitated. A second adhesive alignment ... More
Structure and assembly of the catalytic region of human complement protease C1r: a three-dimensional model based on chemical cross-linking and homology modeling.
AuthorsLacroix M, Rossi V, Gaboriaud C, Chevallier S, Jaquinod M, Thielens NM, Gagnon J, Arlaud GJ
JournalBiochemistry
PubMed ID9174342
C1r is the modular serine protease responsible for autocatalytic activation of C1, the first component of the complement classical pathway. Its catalytic region is a noncovalent homodimer of two gamma-B monomers, each comprising two contiguous complement control protein (CCP) modules, IV and V [also known as short consensus repeats (SCRs)], ... More
Effect of nucleotides and actin on the intramolecular cross-linking of myosin subfragment-1.
AuthorsBlotnick E, Muhlrad A
JournalBiochemistry
PubMed ID8204621
The heavy chain of myosin subfragment-1 (S1) is cleaved by limited trypsinolysis into three fragments, 27, 50, and 20 kDa--aligned in this order from the N-terminus. The tertiary structure of the molecule is essentially not affected by trypsinolysis. The spatial relations between the various regions of the molecule and the ... More
ATP synthase complex. Proximities of subunits in bovine submitochondrial particles.
AuthorsBelogrudov GI, Tomich JM, Hatefi Y
JournalJ Biol Chem
PubMed ID7836433
The catalytic sector, F1, and the membrane sector, F0, of the mitochondrial ATP synthase complex are joined together by a 45-A-long stalk. Knowledge of the composition and structure of the stalk is crucial to investigating the mechanism of conformational energy transfer between F0 and F1. This paper reports on the ... More
Interactions of beta-thymosins, thymosin beta 4-sulfoxide, and N-terminally truncated thymosin beta 4 with actin studied by equilibrium centrifugation, chemical cross-linking and viscometry.
AuthorsHuff T, Zerzawy D, Hannappel E
JournalEur J Biochem
PubMed ID7607239
All beta-thymosins studied interact with G-actin in a bimolecular complex and inhibit the polymerization to F-actin under high salt conditions. The interactions between actin and beta-thymosins have been studied under polymerization conditions using actin labeled by a fluorescent reporter group at Cys374. Instead of labeling actin we employed equilibrium centrifugation ... More
A novel use for a carbodiimide compound for the fixation of fluorescent and non-fluorescent calcium indicators in situ following physiological experiments.
AuthorsTymianski M, Bernstein GM, Abdel-Hamid KM, Sattler R, Velumian A, Carlen PL, Razavi H, Jones OT
JournalCell Calcium
PubMed ID9105727
The inability to determine the precise intracellular location of non-fluorescent organic calcium chelators such as BAPTA is a persistent problem which has precluded much detailed analysis of the chelators' spatial or temporal dynamics in live cells. Similarly, following physiological experiments with fluorescent indicators like Fura-2, it has often been desirable ... More
A facile, water-soluble method for modification of proteins with DOTA. Use of elevated temperature and optimized pH to achieve high specific activity and high chelate stability in radiolabeled immunoconjugates.
AuthorsLewis MR, Raubitschek A, Shively JE
JournalBioconjug Chem
PubMed ID7873659
We have developed a method for attachment of the macrocyclic chelating agent 1,4,7,10-tetraazacyclododecane N,N',N",N"'-tetraacetic acid (DOTA) to proteins by activation of a single carboxyl group with N-hydroxysulfosuccinimide (sulfo-NHS). The sulfo-NHS active ester of DOTA was prepared in a single step using 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC), and DOTA conjugates of cytochrome c and ... More
The interaction of monomeric actin with two binding sites on Acanthamoeba actobindin.
AuthorsBubb MR, Lewis MS, Korn ED
JournalJ Biol Chem
PubMed ID1995634
Actobindin was previously shown to be an 88-residue polypeptide (Mr 9761) with an internal tandem repeat of 33-34 amino acids. Sedimentation equilibrium experiments have confirmed this Mr for native actobindin. Pyreneglyoxal-labeled actobindin had a similar Mr by sedimentation equilibrium analysis and bound to actin in a manner qualitatively similar to ... More
Purification and characterization of subtilisin cleaved actin lacking the segment of residues 43-47 in the DNase I binding loop.
The protease subtilisin has been reported to cleave skeletal muscle G-actin between Met 47 and Gly 48 generating a core fragment of 33 kDa and a small N-terminal peptide, which remains attached to the core fragment [Schwyter, D. Phillips, M., & Reisler, E. (1989) Biochemistry 28, 5889-5895]. However, amino acid ... More
Interaction of adrenodoxin with P4501A1 and its truncated form P450MT2 through different domains: differential modulation of enzyme activities.
AuthorsAnandatheerthavarada HK, Addya S, Mullick J, Avadhani NG
JournalBiochemistry
PubMed ID9454608
Recently we showed that the beta-naphthoflavone-inducible liver mitochondrial P450MT2 consists of two N-terminal truncated forms of the microsomal P4501A1, termed P450MT2a (+5/1A1) and MT2b (+33/1A1) [Addya et al. (1997) J. Cell Biol. 139, 589-599]. In the present study, we demonstrate that intact P4501A1 and the major mitochondrial form, P450MT2b (routinely ... More
Peroxynitrite induces covalent dimerization of epidermal growth factor receptors in A431 epidermoid carcinoma cells.
Authorsvan der Vliet A, Hristova M, Cross CE, Eiserich JP, Goldkorn T
JournalJ Biol Chem
PubMed ID9822654
Irreversible tyrosine modifications by inflammatory oxidants such as peroxynitrite (ONOO-) can affect signal transduction pathways involving tyrosine phosphorylation. The epidermal growth factor receptor (EGFR), a member of the c-ErbB receptor tyrosine kinase family, is involved in regulation of epithelial cell growth and differentiation, and possible modulation of EGFR-dependent signaling by ... More
The importance of carboxyl groups on the lumenal side of the membrane for the function of the Ca(2+)-ATPase of sarcoplasmic reticulum.
AuthorsWebb RJ, Khan YM, East JM, Lee AG
JournalJ Biol Chem
PubMed ID10625635
The conventional model for transport of Ca(2+) by the Ca(2+)-ATPase of skeletal muscle sarcoplasmic reticulum (SR) involves a pair of binding sites for Ca(2+) that change upon phosphorylation of the ATPase from being high affinity and exposed to the cytoplasm to being low affinity and exposed to the lumen. However, ... More
Synthesis of a stable and specific surface plasmon resonance biosensor surface employing covalently immobilized peptide nucleic acids.
AuthorsBurgener M, Sänger M, Candrian U
JournalBioconjug Chem
PubMed ID11087321
Biosensors allow the real-time and label-free observation of biochemical reactions between various ligands including antigen-antibody reactions and nucleic acids hybridizations. In our studies, we used a surface plasmon resonance biosensor to elucidate the hybridization characteristics of a peptide nucleic acid (PNA) ligand immobilized on sensor surfaces either through covalent or ... More
Chemo-mechanical leak formation in human erythrocytes upon exposure to a water-soluble carbodiimide followed by very mild shear stress. II. Chemical modifications involved.
AuthorsThelen P, Deuticke B
JournalBiochim Biophys Acta
PubMed ID3179292
The chemical reactions underlying the chemo-mechanical leak formation in human erythrocytes upon treatment with the carboxyl-modifying reagent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and subsequent minimal shearing, described in the preceding paper (Thelen, P. and Deuticke, B. (1988) Biochim. Biophys. Acta 944, 285-296), are here characterized in more detail. The capacity to form leaks ... More
Sequence-selective biosensor for DNA based on electroactive hybridization indicators.
AuthorsMillan KM, Mikkelsen SR
JournalAnal Chem
PubMed ID8238927
Deoxyribonucleic acid was covalently immobilized onto oxidized glassy carbon electrode surfaces that had been activated using 1-[3-(dimethylamino)-propyl]-3-ethylcarbodimide hydrochloride and N-hydroxysulfosuccinimide. This reaction is selective for immobilization through deoxyguanosine (dG) residues. Immobilized DNA was detected voltammetrically, using tris (2,2'-bipyridyl)cobalt(III) perchlorate and tris (1,10-phenanthroline)cobalt(III) perchlorate (Co(bpy)3(3+) and Co(phen)3(3+). These complexes are reversibly ... More