Super‐Chelators for Advanced Protein Labeling in Living Cells

Live‐cell labeling, super‐resolution microscopy, single‐molecule applications, protein localization, or chemically induced assembly are emerging approaches, which require specific and very small interaction pairs. The minimal disturbance of protein function is essential to derive unbiased insights into cellular processes. Herein, we define a new class of hexavalent N‐nitrilotriacetic acid (hexaNTA) chelators, displaying the highest affinity and stability of all NTA‐based small interaction pairs described so far. Coupled to bright organic fluorophores with fine‐tuned photophysical properties, the super‐chelator probes were delivered into human cells by chemically gated nanopores. These super‐chelators permit kinetic profiling, multiplexed labeling of His₆‐ and His₁₂‐tagged proteins as well as single‐molecule‐based super‐resolution imaging.     

Insight into the Complexation Mode of Bis(nitrilotriacetic acid) (NTA) Ligands with Ni2+ Involved in the Labeling of Histidine‐Tagged Proteins

According to literature reports and our own findings, the binding of new Ni²⁺‐preloaded bis(nitrilotriacetic acid) (NTA) ligands with polyhistidine‐tagged proteins has been found to be accompanied by a one‐ to two‐order‐of‐magnitude increase in affinity, compared to the binding of a single Ni²⁺‐preloaded NTA moiety. In spite of the introduction of a second NTA chelating group, a cooperative effect that is less than the theoretical maximum has been observed. Herein, we present a rational explanation for the observed stability of the ternary complex involving the postulated bis‐NTA–(Ni²⁺)₂ species and multivalent polyhistidine tags. We have found that prior to the formation of the ternary complex, the Ni²⁺‐preloading step of bis‐NTA ligands does not form the expected bis‐NTA–(Ni²⁺)₂ exclusively. Instead of the major formation of bis‐NTA–(Ni²⁺)₂ species, it appears that cyclic discrete 1:1 and 2:2 entities are predominantly formed. It is proposed that these species interact upon ring‐opening with multivalent histidine tags. The occurrence of this phenomena accounts for the overall one‐ to two‐order‐of‐magnitude increase in affinity of ternary complexes involving bis‐NTA ligands.     

Distinct reaction pathways for some classical and nonclassical transition metal dihydrides

Chlorophosphine gold(I) compounds [Au(PR₃)Cl] (R = Et, Ph) have been used as reagents for distinguishing classical from nonclassical structures in a family of dihydrides or dihydrogen complexes of iron, cobalt, rhodium and iridium stabilized by the tripodal polyphosphine PP₃ [PP₃ = P(CH₂CH₂PPh₂)₃]. Novel mixed transition-metal gold hydrides, including the first iron—gold species, are described.     

Hydrophilic and lipophilic iron chelators with the same complexing abilities

A new series of iron chelators with the same coordination sphere as the water-soluble ligand O-trensox, but featuring a variable hydrophilic-lipophilic balance, have been obtained by grafting oxyethylene chains of variable length on a C-pivot tripodal scaffold. The X-ray structure of a ferric complex exhibiting tris(8-hydroxyquinolinate) coordination and solution thermodynamic properties (pK(a) of the ligands, stability constants of the ferric complexes) have been determined. The complexing ability (pFe(III) values) of the ligands are similar to that of O-trensox. Partition coefficients between water and octanol or chloroform have been measured and transport across a membrane has been mimicked ("shuttle process"). The results of biological assays (iron chelation with free ligands or iron nutrition with ferric complexes) could not be correlated with the partition coefficients. These results call into question the role of distribution coefficients (of the ligands and/or complexes) in the biological activities of iron chelators.     

Novel Lanthanide(III) Ternary Complexes with Naphthoyltrifluoroacetone: A Synthetic and Spectroscopic Study

A series of lanthanide complexes with general formula [Ln(NTA)₃X] were prapared [Ln = Y ( a ), Er ( b ), Eu ( c ), NTA = naphthoyltrifluoroacetone, X = H₂O ( 1 ), phen = phenanthroline ( 2 ), bpyO1 = 2, 2′‐bipyridine N‐oxide ( 3 ), and bpyO2 = 2, 2′‐bipyridine‐N,N′‐dioxide ( 4 )]. The crystal structures of [Eu(NTA)₃bpyO2] ( 4b ), [Er(NTA)₃bpyO1] ( 3c ), and [Er(NTA)₃phen] ( 2c ) were determined. X‐ray crystallographic analysis reveals that the complexes are of mononuclear structure with three NTA and one ancillary ligand. The photoluminescence spectra of 3c and 4b exhibit strong characteristic emissions arising from Eu³⁺ central ion due to the efficient sensitization of bpyO1 and bpyO2, respectively.     

The cocrystal of 3‐hydroxy‐2‐naphthoic acid and 4,4′‐bipyridine

4,4′‐Bipyridine cocrystallizes with 3‐hydroxy‐2‐naphthoic acid in a 1:2 ratio to give a centrosymmetric three‐component supra­molecular adduct, namely 3‐hydroxy‐2‐naphthoic acid–4,4′‐bipyridine (2/1), C₁₁H₈O₃·0.5C₁₀H₈N₂, in which 4,4′‐bipyridine is located on an inversion center. The pyridine–carboxylic acid heterosynthon generates an infinite one‐dimensional hydrogen‐bonded chain viaπ–π inter­actions between naphthyl and 4,4′‐bipyridine groups. The one‐dimensional chains are further assembled into a three‐dimensional network by weak C—H⋯π inter­actions between pyridyl and naphthyl rings, and C—H⋯O inter­actions between 3‐hydroxy‐2‐naphthoic acid mol­ecules.     

Iron uptake and toxicity in Caco-2 cells

The differences between the in vitro effects of iron attributed to valence, chelation, and complexation are known in terms of markers of oxidative stress. Few studies, however, describe the effects of iron on general markers of toxicity used in the testing of cell cultures. The aim of the present study was to determine the toxicity and uptake of different salts and iron complexes in the human intestinal cell line, Caco-2.Cells were incubated with 1.5 mM of different species of iron [FeCl₃/nitrilotriacetic acid (NTA) (1:2), FeCl₃/citric acid (1:2), FeCl₃ and FeSO₄] for 22–24 h. Thereafter, toxicological and uptake experiments were performed.The iron uptake, viability (via MTT assay), and membrane stability (via LDH release) of Caco-2 cells incubated with various iron forms differed significantly from untreated controls which showed no detrimental effects on cells and less iron uptake. The lowest signal for cell viability (MTT assay) was found after the incubation of the cells with FeCl₃/citric acid, being significantly different to treatment with FeCl₃, where the highest MTT signal was detected (p=0.002). No differences between the tested iron species could be found regarding cell proliferation (via serial cell counting) and viability using the trypan blue exclusion test. The lowest membrane damage (via LDH release) was registered in cells treated with FeCl₃/citric acid (1:2), whereas the highest LDH release could be found in cells incubated with FeCl₃/NTA (1:2). The highest intracellular iron concentration (measured via GFAAS) was detected after the treatment of Caco-2 cells with FeCl₃ and FeCl₃/NTA (1:2).This study substantiates the importance of the choice of complexes, as NTA seemed to enhance the toxicity of iron, while citric acid inhibited iron uptake and toxicity.     

Diacetonalkohol‐Komplexe von Lanthanoid‐Trichloriden. Die Kristallstrukturen von [LnCl3(DAA)2] mit Ln = Sm und Eu

Diacetone Alcohol Complexes of Lanthanide Trichlorides. Crystal Structures of [LnCl₃(DAA)₂] with Ln = Sm and Eu The diacetone alcohol complexes [LnCl₃(DAA)₂] with Ln = samarium ( 1 ) and europium ( 2 ) are obtained from the waterfree metal trichlorides with excess diacetone alcohol (4‐hydroxy‐4‐methyl‐2‐pentanone = DAA) forming colourless ( 1 ) and pale yellow crystals ( 2 ), respectively, which are characterized by crystal structure determinations. The europium compound 2 is additionally described by its vibrational spectra (IR, Raman). 1 and 2 crystallize isotypically with one another. The metal atoms of the molecular complex units are unusually coordinated in a distorted pentagonal‐bipyramdial fashion by the three chlorine atoms and by the two alcoholic oxygen atoms of the DAA molecules in the equatorial plane. The apical positions are occupied by the carbonyl oxygen atoms of the chelating DAA molecules. The complex units [LnCl₃(DAA)₂] are associated along [100] by bifurcated —OH···Cl···HO— bridges to form chains. 1 : Space group P2₁, Z = 2, lattice dimensions at —80 °C: : a = 710.2(1), b = 1617.6(2), c = 827.3(1) pm; β = 106.36(1)°; R₁ = 0.026. 2 : Space group P2₁, Z = 2, lattice dimensions at —80 °C: a = 709.7(1), b = 1614.5(2), c = 825.7(1) pm; β = 106.40(1)°; R₁ = 0.0303.     

Intermolecular interactions in the chiral and racemic forms of 3‐­hydroxy‐2‐(1‐oxoisoindolin‐2‐yl)­butanoic acid derived from threonine

The title compounds, C₁₂H₁₃NO₄, are derived from l ‐threonine and dl ‐threonine, respectively. Hydro­gen bonding in the chiral derivative, (2S/3R)‐3‐hydroxy‐2‐(1‐oxoisoindolin‐2‐yl)­butanoic acid, consists of O—H_acid?O_alkyl—H?O=C_indole chains [O?O 2.659 (3) and 2.718 (3) Å], Csp³—H?O and three C—H?π_arene interactions. In the (2R,3S/2S,3R) racemate, conventional carboxylic acid hydrogen bonding as cyclical (O—H?O=C)₂ [graph set R₂²(8)] is present, with O_alkyl—H?O=C_indole, Csp³—H?O and C—H?π_arene interactions. The COOH group geometry differs between the two forms, with C—O, C=O, C—C—O and C—C=O bond lengths and angles of 1.322 (3) and 1.193 (3) Å, and 109.7 (2) and 125.4 (3)°, respectively, in the chiral structure, and 1.2961 (17) and 1.2210 (18) Å, and 113.29 (12) and 122.63 (13)°, respectively, in the racemate structure. The O—C=O angles of 124.9 (3) and 124.05 (14)° are similar. The differences arise from the contrasting COOH hydrogen‐bonding environments in the two structures.     

New pharmaceutical salts containing pyridoxine

Two mixed crystals were obtained by crystallizing the active pharmaceutical ingredient pyridoxine [systematic name: 4,5‐bis(hydroxymethyl)‐2‐methylpyridin‐3‐ol, PN] with (E )‐3‐(4‐hydroxy‐3‐methoxyphenyl)prop‐2‐enoic acid (ferulic acid) and 4‐hydroxy‐3,5‐dimethoxybenzoic acid (syringic acid). PN and the coformers crystallize in the form of pharmaceutical salts in a 1:1 stoichiometric ratio, namely 3‐hydroxy‐4,5‐bis(hydroxymethyl)‐2‐methylpyridin‐1‐ium (E )‐3‐(4‐hydroxy‐3‐methoxyphenyl)prop‐2‐enoate, C₈H₁₂NO₃⁺·C₉H₉O₅⁻, and 3‐hydroxy‐4,5‐bis(hydroxymethyl)‐2‐methylpyridin‐1‐ium 4‐hydroxy‐3,5‐dimethoxybenzoate monohydrate, C₈H₁₂NO₃⁺·C₁₀H₁₁O₅⁻·H₂O, the proton exchange between PN and the acidic partner being supported by the differences of the pK _a values of the two components and by the C—O bond lengths of the carboxylate groups. Besides complex hydrogen‐bonding networks, π–π interactions between aromatic moieties have been found to be important for the packing architecture in both crystals. Hirshfeld surface analysis was used to explore the intermolecular interactions in detail and compare them with the interactions found in similar pyridoxine/carboxylic acid salts.     

Iron chelating agents for the treatment of iron overload

The importance of iron chelators in medicine has significantly increased in recent years. Iron is essential for life but it is also potentially more toxic than other trace elements. This is because we lack effective means to protect human cells against iron overload and because of the role of iron in the generation of free radicals. In order to protect patients from the consequences of iron toxicity, iron chelating agents have been introduced in clinical practice. Unfortunately, the ideal chelator for treating iron overload in humans has not been identified yet. In this paper we examine a few characteristics of iron chelators, with some emphasis on the effects of redox cycling, on absorption mechanisms and on some properties of the pFe. A brief summary is then made of the chelators recently proposed or in development for the treatment of iron overload.     

Synthesis and Crystal Structure of Novel Coordination Polymers with Nitrilotripropionic Acid

A novel three‐dimensional (3D) lanthanide‐organic framework [Pr₃(NTP)₃(H₂O)₆] · 10H₂O ( 1 ), (H₃NTP = 3,3′,3′‐nitrilotripropionic acid) was synthesized and characterized by elemental analyses, IR spectroscopy, and X‐ray diffraction analyses. The results show that complex 1 is connected through NTP ligands to form a 3D network with microchannels. The coordination mode of the NTP ligand was found for the first time. In order to investigate the temperature effects on controlling the dimensionality of the complexes, another two complexes, namely, [Nd(NTP)(H₂O)₂] · H₂O · HBr ( 2 ) and [Pr(H₂O)][⁺NH₃(CH₂CH₂COO^–)][OOCCOO]_1.5 ( 3 ) were synthesized and characterized. Notably, in complex 3 , the NTP ligand lost its two arms because of the high temperature. Furthermore, the thermogravimetric analyses of the three complexes are discussed in detail.     

Facile C=O Bond Splitting of Carbon Dioxide Induced by Metal–Ligand Cooperativity in a Phosphinine Iron(0) Complex

New iron complexes [Cp*Fe L ]^? ( 1‐σ and 1‐π , Cp*=C₅Me₅) containing the chelating phosphinine ligand 2‐(2′‐pyridyl)‐4,6‐diphenylphosphinine ( L ) have been prepared, and found to undergo facile reaction with CO₂ under ambient conditions. The outcome of this reaction depends on the coordination mode of the versatile ligand L . Interaction of CO₂ with the isomer 1‐π , in which L binds to Fe through the phosphinine moiety in an η⁵ fashion, leads to the formation of 3‐π , in which CO₂ has undergone electrophilic addition to the phosphinine group. In contrast, interaction with 1‐σ —in which L acts as a σ‐chelating [P,N] ligand—leads to product 3‐σ in which one C=O bond has been completely broken. Such CO₂ cleavage reactions are extremely rare for late 3d metals, and this represents the first such example mediated by a single Fe centre.     

Speciation Studies of Iron (III) with (Poly)phosphonate Ligands: The Case of EDTMP (EthylenediamineTetramethylene Phosphonic Acid)

The complex formation equilibrium of ethylenediaminetetramethylenephosphonic acid (EDTMP, H₈L) with iron (III) has been studied potentiometrically at 25°C and an ionic strength of 0.2 M (NaCl). The successive protonation constants of ligand EDTMP and the complex formation constants were determined with the PSEQUAD program. Keeping in view the biological studies, the speciation in the system Fe (III)—EDTMP was calculated and drawn with the HySS computer program, and pFe values are compared.     

Two succinic acid derivatives of 2‐ethyl‐6‐methylpyridin‐3‐ol

Crystals of bis(2‐ethyl‐3‐hydroxy‐6‐methylpyridinium) succinate–succinic acid (1/1), C₈H₁₂NO⁺·0.5C₄H₄O₄²⁻·0.5C₄H₆O₄, (I), and 2‐ethyl‐3‐hydroxy‐6‐methylpyridinium hydrogen succinate, C₈H₁₂NO⁺·C₄H₅O₄⁻, (II), were obtained by reaction of 2‐ethyl‐6‐methylpyridin‐3‐ol with succinic acid. The succinate anion and succinic acid molecule in (I) are located about centres of inversion. Intermolecular O—H...O, N—H...O and C—H...O hydrogen bonds are responsible for the formation of a three‐dimensional network in the crystal structure of (I) and a two‐dimensional network in the crystal structure of (II). Both structures are additionally stabilized by π–π interactions between symmetry‐related pyridine rings, forming a rod‐like cationic arrangement for (I) and cationic dimers for (II).     

The epimeric 9‐oxobicyclo[3.3.1]nonane‐3‐carboxylic acids: hydrogen‐bonding patterns of the endo acid and the lactol of the exo acid

The two δ‐keto carboxylic acids of the title, both C₁₀H₁₄O₃, are epimeric at the site of carboxyl attachment. The endo (3α) epimer, (I), has its keto‐acid ring in a boat conformation, with the tilt of the carboxyl group creating conformational chirality. The mol­ecules form hydrogen bonds by centrosymmetric pairing of carboxyl groups across the corners of the chosen cell [O⃛O = 2.671 (2) Å and O—H⃛O = 179 (2)°]. Two close intermolecular C—H⃛O contacts exist for the ketone. The exo (3β) epimer exists in the closed ring–chain tautomeric form as the lactol, 8‐hydroxy‐9‐oxatri­cyclo­[5.3.1.0^3,8]­undecan‐10‐one, (II). The mol­ecules have conformational chirality, and the hydrogen‐bonding scheme involves intermolecular hydroxyl‐to‐carbonyl chains of mol­ecules screw‐related in b [O⃛O = 2.741 (2) Å and O—H⃛O = 177 (2)°].     

μ‐Oxo‐bis{[(6‐hydroxy­methyl‐2‐pyridyl­methyl)­bis(2‐pyridyl­methyl)­amine‐κ5N,N′,N′′,N′′′,O]­iron(III)} tetrakis­(perchlorate)

The novel μ‐oxo‐diiron complex [Fe₂O(BPHPA)₂](ClO₄)₄ [BPHPA is (6‐hydroxy­methyl‐2‐pyridyl­methyl)­bis(2‐pyridyl­methyl)­amine, C₁₉H₂₀N₄O], contains a binuclear centrosymmetric [Fe₂O(BPHPA)₂]⁴⁺ cation (the bridging O atom lies on an inversion centre) and four perchlorate anions. Each iron ion is coordinated by four N atoms [Fe—N = 2.117 (5)–2.196 (5) Å] and one O atom [Fe—O = 2.052 (5) Å] from a BPHPA ligand, and by one bridging oxo atom [Fe—O = 1.7896 (9) Å], forming a distorted octahedron. There are hydrogen bonds between the hydroxy group and perchlorate O atoms [O—H·O = 2.654 (7) Å].     

Solution Properties of Azidokojatoiron(III) Complexes

The formation of iron(III) complexes with chelating azidokojate anions L⁻ was investigated in aqueous solutions as a function of the pH and the c(Fe³⁺):c(HL) molar ratio. Based on the stability constants, the distribution among the above complexes, [Fe(H₂O)₆]³⁺, and [Fe(H₂O)₅(OH)]²⁺ were calculated in solutions of various compositions. The complexes are redox stable in aqueous solutions both in the dark and in visible laboratory light. Properties of the investigated azidokojic acid and its iron(III) complexes are compared with those required for therapeutic applications as alternative iron chelators.     

Aqueous polymerization of acrylamide initiated by cerium(IV)–amino acid chelating agent redox initiators

Amino acid-type chelating agents such as nitrilotriacetic acid (NTA), nitrilotripropionic acid (NPA), iminodiacetic acid (IDA), and ethylenediamine tetraacetic acid (EDTA) were used in combination with cerium(IV) ammonium nitrate [Ce(IV)] as the redox initiators for the aqueous polymerizations of acrylamide (AM). The polymerization behaviors and polymer qualities were studied as functions of the concentrations of Ce(IV), chelating agent, AM, as well as temperature. The performances of the chelating agent redox systems varied with the natures of the chelating agents. The NTA–Ce(IV) initiator showed the most promising polymerization rate and conversion. The blank tests for the reactions of cerium and chelating agents were also conducted for finding mechanism of formation of free radicals and determining their complex formation constants (K) and disproportionation constants (k_d). The mechanism for the polymerization was proposed and the kinetic parameters were evaluated. © 1993 John Wiley & Sons, Inc.     

Syntheses and Structural Characterization of Two Nitrilotriacetate Cobalt Complexes: [{CoK2(NTA)(Hmta)(H2O)3}NO3]n and [{Co(4,4′‐bpy)2(H2O)4}{Co2(NTA)2(4,4′‐bpy)(H2O)2}]

Two nitrilotriacetate cobalt complexes {[CoK₂(NTA)(Hmta)(H₂O)₃]NO₃}_n ( 1 ) and [{Co(4,4′‐bpy)₂(H₂O)₄}{Co₂(NTA)₂(4,4′‐bpy)(H₂O)₂}] ( 2 ) (NTA = nitrilotriacetate anion, Hmta = hexamethylenetetramine and 4,4′‐bpy = 4,4′‐bipyridine) were prepared and characterized by IR, elemental analysis and single crystal X‐ray diffraction study. The influence of the neutral ancillary ligands on the formation of the complexes with different structures in the Co‐NTA system was discussed. The coordination of NTA and Hmta to Co²⁺ ions only resulted in the formation of mononuclear [Co(NTA)(Hmta)]^? ions which are further connected by K⁺ ions and water molecules to form a three‐dimensional network. The use of 4,4′‐bpy as ancillary ligand in 2 led to the formation of separate mononuclear [Co(4,4′‐bpy)₂(H₂O)₄]²⁺ and dinuclear [Co₂(NTA)₂(4,4′‐bpy)(H₂O)₂]^2? which are further connected by hydrogen bonds to form a supramolecular three‐dimensional network. In these cases it seems to suggest that the addition of neutral ancillary ligand into the Co‐NTA system leads to the formation of lower dimensional structures when the contribution of alkali ions to the structural dimensionality is neglected.