Influence of Chemical Modifications of the Crystallophore on Protein Nucleating Properties and Supramolecular Interactions Network.

Crystallophores are lanthanide complexes that have demonstrated outstanding induction of crystallization for various proteins. This article explores the effect of tailored modifications of the crystallophore first generation and their impact on the nucleating properties and protein crystal structures. Through high-throughput crystallization experiments and dataset analysis, we evaluated the effectiveness of these variants, in comparison to the first crystallophore generation G1. In particular, the V1 variant, featuring a propanol pendant arm, demonstrated the ability to produce new crystallization conditions for the proteins tested (hen-egg white lysozyme, proteinase K and thaumatin). Structural analysis performed in the case of hen egg-white lysozyme along with Molecular Dynamics simulations, highlights V1's unique behavior, taking advantage of the flexibility of its propanol arm to explore different protein surfaces and form versatile supramolecular interactions.

One touch is all it takes: the supramolecular interaction between ubiquitin and lanthanide complexes revisited by paramagnetic NMR and molecular dynamics.

The supramolecular interaction between lanthanide complexes and proteins is at the heart of numerous chemical and biological studies. Some of these complexes have demonstrated remarkable interaction properties with proteins or peptides in solution and in the crystalline state. Here we have used the paramagnetism of lanthanide ions to characterize the affinity of two lanthanide complexes for ubiquitin. As the interaction process is dynamic, the acquired NMR data only reflect the time average of the different steps. We have used molecular dynamics (MD) simulations to get a deeper insight into the detailed interaction scenario at the microsecond scale. This NMR/MD approach enabled us to establish that the tris-dipicolinate complex interacts specifically with arginines and lysines, while the crystallophore explores the protein surface through weak interactions with carboxylates. These observations shed new light on the dynamic interaction properties of these complexes, which will ultimately enable us to propose a crystallization mechanism.

Temperature-dependent NIR-CPL spectra of chiral Yb(III) complexes.

Chiral, enantiopure Yb(III) complexes exhibit circularly polarized luminescence (CPL) in the near infrared (NIR) wavelength region. This CPL is quantified by the dissymmetry factor (glum). The excited state 2F5/2 consists of six mJ' states degenerated in three Stark levels, due to the crystal-field splitting (CFS), which are populated in accordance with the Boltzmann distribution. Consequently, room temperature CPL spectra are the sum of various - either positive or negative - contributions, that are practically impossible to quantify. To address this issue, an advanced setup enabling CPL measurements over a broad temperature range (300 to 4 K) has been developed. The interrelation of CFS, glum and temperature was explored using a pair of enantiopure Yb(III) complexes, highlighting the individual contribution of each crystal-field sublevel to the overall CPL spectrum, as anticipated by simulations performed in the framework of multireference wave-functions. Hence, the CPL spectra of chiral lanthanide complexes were found to be indeed strongly temperature-dependent, as is the glum dissymmetry factor, as a consequence of the variation in thermal sublevel population.

Ytterbium(III) Complex with Photochromic Ruthenium(II) Acetylide Ligand: All Visible Light Photoswitching of NIR Luminescence.

We report a ruthenium(II) bisacetylide complex bearing a photochromic dithienylethene (DTE) acetylide arm and a coordinating bipyridyl on the trans acetylide unit. Its coordination with Yb(TTA)3 centers (TTA = 2-thenoyltrifluoroacetonate) produces a bimetallic complex in which the dithienylethene isomerization is triggered by both ultraviolet (UV) light absorbed by the DTE unit and 450 nm excitation in a transition of the organometallic moiety. The redox behavior arising from the ruthenium(II) bisacetylide system is fully investigated by cyclic voltammetry and spectroelectrochemistry, revealing a lack of stability of the DTE-closed oxidized state preventing effective redox luminescence switching. On the other hand, the photoswitching of ytterbium(III) near-infrared (NIR) emission triggered by the photochromic reaction is fully operational. The electronic structure of this complex in its different states characterized by strong electronic coupling between the DTE and the ruthenium(II)-based moieties leading to metal-assisted photochromic behavior were rationalized with the help of time-dependent density functional theory (TD-DFT) calculations.

Distributive Nd-to-Yb Energy Transfer within Pure [YbNdYb] Heterometallic Molecules.

Facile access to site-selective hetero-lanthanide molecules will open new avenues in the search of novel photophysical phenomena based on Ln-to-Ln' energy transfer (ET). This challenge demands strategies to segregate efficiently different Ln metal ions among different positions in a molecule. We report here the one-step synthesis and structure of a pure [YbNdYb] (1) coordination complex featuring short Yb···Nd distances, ideal to investigate a potential distributive (i.e., from one donor to two acceptors) intramolecular ET from one Nd3+ ion to two Yb3+ centers within a well-characterized molecule. The difference in ionic radius is the mechanism allowing to allocate selectively both types of metal ion within the molecular structure, exploited with the simultaneous use of two ß-diketone-type ligands. To assist the photophysical investigation of this heterometallic species, the analogues [YbLaYb] (2) and [LuNdLu] (3) have also been prepared. Sensitization of Yb3+ and Nd3+ in the last two complexes, respectively, was observed, with remarkably long decay times, facilitating the determination of the Nd-to-Yb ET within the [YbNdYb] composite. This ET was demonstrated by comparing the emission of iso-absorbant solutions of 1, 2, and 3 and through lifetime determinations in solution and solid state. The comparatively high efficiency of this process corroborates the facilitating effect of having two acceptors for the nonradiative decay of Nd3+ created within the [YbNdYb] molecule.

Multifunctional Helicene-Based Ytterbium Coordination Polymer Displaying Circularly Polarized Luminescence, Slow Magnetic Relaxation and Room Temperature Magneto-Chiral Dichroism.

The combination of physical properties sensitive to molecular chirality in a single system allows the observation of fascinating phenomena such as magneto-chiral dichroism (MChD) and circularly polarized luminescence (CPL) having potential applications for optical data readout and display technology. Homochiral monodimensional coordination polymers of YbIII were designed from a 2,15-bis-ethynyl-hexahelicenic scaffold decorated with two terminal 4-pyridyl units. Thanks to the coordination of the chiral organic chromophore to Yb(hfac)3 units (hfac- =1,1,1,5,5,5-hexafluoroacetylaconate), efficient NIR-CPL activity is observed. Moreover, the specific crystal field around the YbIII induces a strong magnetic anisotropy which leads to a single-molecule magnet (SMM) behaviour and a remarkable room temperature MChD. The MChD-structural correlation is supported by computational investigations.

Synthesis and Photophysical Properties of 1,1,4,4-Tetracyanobutadienes Derived from Ynamides Bearing Fluorophores.

1,1,4,4-Tetracyanobutadienes (TCBDs) bearing a large diversity of fluorophores were prepared following a multi-step synthesis. In a crucial last step, all compounds were obtained from the corresponding ynamides, which were particularly suitable for the formation of the TCBDs in the presence of tetracyanoethylene via a [2+2] cycloaddition/retroelectrocyclization step (CA-RE). Several fluorenyl derivatives in addition to phenanthrenyl and terphenyl ones provided ynamide-based TCBDs affording remarkable emission properties covering a large range of wavelengths. Those compounds emit both in solid state and in solution from the visible region to the NIR range, depending on the molecular structures. Quantum yields in cyclohexane reached unforeseen values for such derivatives, up to 7.8 %. A huge sensitivity to the environment of the TCBDs has also been unraveled for most of the compounds since we observed a dramatic fall of the quantum yields when changing the solvent from cyclohexane to toluene, while they are almost non-emissive in dichloromethane.

Circularly polarized luminescence of Eu(III) complexes with chiral 1,1'-bi-2-naphtol-derived bisphosphate ligands.

The interest for lanthanide circularly polarized luminescence (CPL) has been quickly growing for 10 years. However, very few of these studies have involved correlation between the dissymmetry factor (glum ) and the chemical modifications in a series of chiral ligands. Four polymeric compounds of Eu(III) were prepared by using a series of binaphtyl derivatives for which the size of the π system as well as the number of stereogenic elements (i.e., the binaphtyl moiety) are modulated. The resulting {[Eu(hfac)3 ((S)/(R)-Lx )]}n (x = 1 and 3) and {[Eu(hfac)3 ((S,S,S)/(R,R,R)-Lx )]}n (x = 2 and 4) have been characterized by powder X-ray diffraction by comparison with the X-ray structures on single crystal of the Dy(III) analogs. In solution, the structure of the complexes is deeply modified and becomes monomeric. The nature of the ligand induces change in the shape of the CPL spectra in CH2 Cl2 solution. Furthermore, a large |glum | = 0.12 of the magnetic-dipole transition for the [Eu(hfac)3 ((S,S,S)/(R,R,R)-L2 )] complex involving the ligand with three stereogenic elements and an extended 𝜋 system has been measured. This report also shows CPL measurements in solid state for the series of {[Eu(hfac)3 ((S)/(R)-Lx )]}n (x = 1 and 3) and {[Eu(hfac)3 ((S,S,S)/(R,R,R)-Lx )]}n (x = 2 and 4) polymers.

Solid-State Near-Infrared Circularly Polarized Luminescence from Chiral YbIII -Single-Molecule Magnet.

A field-induced chiral YbIII Single-Molecule Magnet (SMM) displayed an unprecedented near-infrared circularly polarized luminescence (NIR-CPL) in the solid-state. The bridging bis(1,10-phenantro[5,6b])tetrathiafulvalene triad (L) allowed an efficient sensitization of the NIR 2 F5/2 →2 F7/2 emission while the NIR-CPL is associated to the f-f transitions of the YbIII ion bearing chiral ß-diketonate derived-camphorate ancillary ligands.

Tuning Excited-State Properties of [2.2]Paracyclophane-Based Antennas to Ensure Efficient Sensitization of Lanthanide Ions or Singlet Oxygen Generation.

The multistep synthesis of original antennas incorporating substituted [2.2]paracyclophane (pCp) moieties in the π-conjugated skeleton is described. These antennas, functionalized with an electron donor alkoxy fragment (A1) or with a fused coumarin derivative (A2), are incorporated in a triazacyclonane macrocyclic ligand L1 or L2, respectively, for the design of Eu(III), Yb(III), and Gd(III) complexes. A combined photophysical/theoretical study reveals that A1 presents a charge transfer character via through-space paracyclophane conjugation, whereas A2 presents only local excited states centered on the coumarin-paracyclophane moiety, strongly favoring triplet state population via intersystem crossing. The resulting complexes EuL1 and YbL2 are fully emissive in red and near-infrared, respectively, whereas the GdL2 complex acts as a photosensitizer for the generation of singlet oxygen.

Influence of Divalent Cations in the Protein Crystallization Process Assisted by Lanthanide-Based Additives.

The use of lanthanide complexes as powerful auxiliaries for biocrystallography prompted us to systematically analyze the influence of the commercial crystallization kit composition on the efficiency of two lanthanide additives: [Eu(DPA)3]3- and Tb-Xo4. This study reveals that the tris(dipicolinate) complex presents a lower chemical stability and a strong tendency toward false positives, which are detrimental for its use in a high-throughput robotized crystallization platform. In particular, the crystal structures of (Mg(H2O)6)3[Eu(DPA)3]2·7H2O (1), {(Ca(H2O)4)3[Eu(DPA)3]2}n·10nH2O (2), and {Cu(DPA)(H2O)2}n (3), resulting from spontaneous crystallization in the presence of a divalent alkaline-earth cation and transmetalation, are reported. On the other hand, Tb-Xo4 is perfectly soluble in the crystallization media, stable in the presence of alkaline-earth dications, and slowly decomposes (within days) by transmetalation with transition metals. The original structure of [Tb4L4(H2O)4]Cl4·15H2O (4) is also described, where L represents a bis(pinacolato)triazacyclononane ligand. This paper also highlights a potential synergy of interactions between Tb-Xo4 and components of the crystallization mixtures, leading to the formation of complex adducts like {AdkA/Tb-Xo4/Mg2+/glycerol} in the protein binding sites. The observation of such multicomponent adducts illustrated the complexity and versatility of the supramolecular chemistry occurring at the surface of the proteins.

Capturing the dynamic association between a tris-dipicolinate lanthanide complex and a decapeptide: a combined paramagnetic NMR and molecular dynamics exploration.

In the realm of biomolecules, peptides can present a large diversity of structures. Our study sheds new light on the structural interplay between a tris-dipicolinate lanthanide probe and a decapeptide SASYKTLPRG. Although a rather trivial, electrostatically driven interaction was expected, the combination of paramagnetic NMR and molecular dynamics simulations reveals a highly dynamic association process and allows for providing extensive insights into the interaction sites and their occupancy. This study highlights the importance of a large conformational sampling to reconcile characteristic time in NMR with molecular dynamics simulations, where sampling in the microsecond range is needed. This study opens the door for a detailed mechanistic elucidation of the early steps of lanthanide complex-peptide or lanthanide complex-protein interaction or self-assembly processes.

Archaeal acetoacetyl-CoA thiolase/HMG-CoA synthase complex channels the intermediate via a fused CoA-binding site.

Many reactions within a cell are thermodynamically unfavorable. To efficiently run some of those endergonic reactions, nature evolved intermediate-channeling enzyme complexes, in which the products of the first endergonic reactions are immediately consumed by the second exergonic reactions. Based on this concept, we studied how archaea overcome the unfavorable first reaction of isoprenoid biosynthesis-the condensation of two molecules of acetyl-CoA to acetoacetyl-CoA catalyzed by acetoacetyl-CoA thiolases (thiolases). We natively isolated an enzyme complex comprising the thiolase and 3-hydroxy-3-methylglutaryl (HMG)-CoA synthase (HMGCS) from a fast-growing methanogenic archaeon, Methanothermococcus thermolithotrophicus HMGCS catalyzes the second reaction in the mevalonate pathway-the exergonic condensation of acetoacetyl-CoA and acetyl-CoA to HMG-CoA. The 380-kDa crystal structure revealed that both enzymes are held together by a third protein (DUF35) with so-far-unknown function. The active-site clefts of thiolase and HMGCS form a fused CoA-binding site, which allows for efficient coupling of the endergonic thiolase reaction with the exergonic HMGCS reaction. The tripartite complex is found in almost all archaeal genomes and in some bacterial ones. In addition, the DUF35 proteins are also important for polyhydroxyalkanoate (PHA) biosynthesis, most probably by functioning as a scaffold protein that connects thiolase with 3-ketoacyl-CoA reductase. This natural and highly conserved enzyme complex offers great potential to improve isoprenoid and PHA biosynthesis in biotechnologically relevant organisms.

Luminescence-Driven Electronic Structure Determination in a Textbook Dimeric DyIII -Based Single-Molecule Magnet.

A textbook dysprosium dinuclear complex based on acetylacetone ligands, [Dy2 (acac)4 (µ2 -acac)2 (H2 O)2 ], has been synthesized and fully characterized. This simple dimeric lanthanide complex shows well-resolved solid-state luminescence and behaves as a single-molecule magnet under zero DC field. A seminal crystal-field approach is used to marry both magnetism and luminescence in the frame of an energetic picture.

The multicatalytic compartment of propionyl-CoA synthase sequesters a toxic metabolite.

Cells must cope with toxic or reactive intermediates formed during metabolism. One coping strategy is to sequester reactions that produce such intermediates within specialized compartments or tunnels connecting different active sites. Here, we show that propionyl-CoA synthase (PCS), an ∼ 400-kDa homodimer, three-domain fusion protein and the key enzyme of the 3-hydroxypropionate bi-cycle for CO2 fixation, sequesters its reactive intermediate acrylyl-CoA. Structural analysis showed that PCS forms a multicatalytic reaction chamber. Kinetic analysis suggested that access to the reaction chamber and catalysis are synchronized by interdomain communication. The reaction chamber of PCS features three active sites and has a volume of only 33 nm3. As one of the smallest multireaction chambers described in biology, PCS may inspire the engineering of a new class of dynamically regulated nanoreactors.

Cationic Biphotonic Lanthanide Luminescent Bioprobes Based on Functionalized Cross-Bridged Cyclam Macrocycles.

Cationic lanthanide complexes are generally able to spontaneously internalize into living cells. Following our previous works based on a diMe-cyclen framework, a second generation of cationic water-soluble lanthanide complexes based on a constrained cross-bridged cyclam macrocycle functionalized with donor-π-conjugated picolinate antennas was prepared with europium(III) and ytterbium(III). Their spectroscopic properties were thoroughly investigated in various solvents and rationalized with the help of DFT calculations. A significant improvement was observed in the case of the Eu3+ complex, while the Yb3+ analogue conserved photophysical properties in aqueous solvent. Two-photon (2P) microscopy imaging experiments on living T24 human cancer cells confirmed the spontaneous internalization of the probes and images with good signal-to-noise ratio were obtained in the classic NIR-to-visible configuration with the Eu3+ luminescent bioprobe and in the NIR-to-NIR with the Yb3+ one.

Redox-Modulations of Photophysical and Single-molecule Magnet Properties in Ytterbium Complexes Involving Extended-TTF Triads.

The reaction between the 2,2'-benzene-1,4-diylbis(6-hydroxy-4,7-di-tert-butyl-1,3-benzodithiol-2-ylium-5-olate triad (H2SQ) and the metallo-precursor [Yb(hfac)3]2H2O led to the formation of a dinuclear coordination complex of formula [Yb2(hfac)6(H2SQ)]0.5CH2Cl2 (H2SQ-Yb). After chemical oxidation of H2SQ in 2,2'-cyclohexa-2,5-diene-1,4-diylidenebis(4,7-di-tert-butyl-1,3-benzodithiole-5,6-dione (Q), the latter triad reacted with the [Yb(hfac)3]2H2O precursor to give the dinuclear complex of formula [Yb2(hfac)6(Q)] (Q-Yb). Both dinuclear compounds have been characterized by X-ray diffraction, DFT optimized structure and electronic absorption spectra. They behaved as field-induced Single-Molecule Magnets (SMMs) nevertheless the chemical oxidation of the semiquinone to quinone moieties accelerated by a factor of five the relaxation time of the magnetization of Q-Yb compared to the one for H2SQ-Yb. The H2SQ triad efficiently sensitized the YbIII luminescence while the chemical oxidation of H2SQ into Q induced strong modification of the absorption properties and thus a quenching of the YbIII luminescence for Q-Yb. In other words, both magnetic modulation and luminescence quenching are reached by the oxidation of the protonated semiquinone into quinone.

Dual Light and Redox Control of NIR Luminescence with Complementary Photochromic and Organometallic Antennae.

With the help of a judicious association between dithienylethene (DTE) units, an ytterbium ion, and a ruthenium carbon-rich complex, we describe (i) the efficient (on/off) switching of pure NIR luminescence with a photochromic unit absorbing in the UV range and (ii) the association of electrochemical and photochemical control of this NIR emission in a single system with nondestructive readout.

Intriguing Effects of Halogen Substitution on the Photophysical Properties of 2,9-(Bis)halo-Substituted Phenanthrolinecopper(I) Complexes.

Three new copper(I) complexes [Cu(LX)2]+(PF6-) (where LX stands for 2,9-dihalo-1,10-phenanthroline and X = Cl, Br, and I) have been synthesized in order to study the impact of halogen substituents tethered in the α position of the chelating nitrogen atoms on their physical properties. The photophysical properties of these new complexes (hereafter named Cu-X) were characterized in both their ground and excited states. Femtosecond ultrafast spectroscopy revealed that early photoinduced processes are faster for Cu-I than for Cu-Cl or Cu-Br, both showing similar behaviors. Their electronic absorption and electrochemical properties are comparable to benchmark [Cu(dmp)2]+ (where dmp stands for 2,9-dimethyl-1,10-phenanthroline); furthermore, their optical features were fully reproduced by time-dependent density functional theory and ab initio molecular dynamics calculations. All three complexes are luminescent at room temperature, showing that halogen atoms bound to positions 2 and 9 of phenanthroline are sufficiently bulky to prevent strong interactions between the excited Cu complexes and solvent molecules in the coordination sphere. Their behavior in the excited state, more specifically the extent of the photoluminescence efficiency and its dependence on the temperature, is, however, strongly dependent on the nature of the halogen. A combination of ultrafast transient absorption spectroscopy, temperature-dependent steady-state fluorescence spectroscopy, and computational chemistry allows one to gain a deeper understanding of the behavior of all three complexes in their excited state.

Unveiling the Binding Modes of the Crystallophore, a Terbium-based Nucleating and Phasing Molecular Agent for Protein Crystallography.

Crystallophores are lanthanide complexes that act as powerful auxiliary for protein crystallography due to their strong nucleating and phasing effects. To get first insights on the mechanisms behind nucleation induced by Crystallophore, we systematically identified various elaborated networks of supramolecular interactions between Tb-Xo4 and subset of 6 protein structures determined by X-ray diffraction in complex with terbium-Crystallophore (Tb-Xo4). Such interaction mapping analyses demonstrate the versatile binding behavior of the Crystallophore and pave the way to a better understanding of its unique properties.