Structure of deoxyhaemoglobin Yakima: a high-affinity mutant form exhibiting oxy-like 1 2 subunit interactions

Crystals of deoxyhaemoglobin Yakima (Asp Gl(99)β → His) are isomorphous with those of deoxyhaemoglobin A, even though the mutation produces disturbances in both the tertiary structure of the subunits and the quaternary structure of the tetramer. Asp Gl(99)β₂ lies at the α₁β₂ subunit interface, and in deoxyhaemoglobin A forms a crucial hydrogen bond with Tyr C7(42) α₁. The histidine residue that replaces the aspartate results in the removal of this single important intersubunit bond, and it further acts as a wedge between the α₁ and β₂ subunits, so that they are pushed apart and displaced part of the way towards the oxy structure. These disturbances are accompanied by the formation of a new intersubunit hydrogen bond, which is usually only observed in the oxy quaternary structure of haemoglobin. The disturbances at the α₁β₂ contact affect the stereochemistry of the entire molecule and are transmitted to the α and β haems. The X-ray structure of deoxy Yakima therefore provides a stereochemical explanation for its abnormal function; this being an abnormally high affinity for oxygen and vastly diminished haem-haem interactions.     

Difference in the structures of alanine tri‐ and tetra‐peptides with antiparallel β‐sheet assessed by X‐ray diffraction,solid‐state NMR and chemical shift calculations by GIPAW

Alanine oligomers provide a key structure for silk fibers from spider and wild silkworms.We report on structural analysis of l ‐alanyl‐l ‐alanyl‐l ‐alanyl‐l ‐alanine (Ala)₄ with anti‐parallel (AP) β‐structures using X‐ray and solid‐state NMR. All of the Ala residues in the (Ala)₄ are in equivalent positions, whereas for alanine trimer (Ala)₃ there are two alternative locations in a unit cell as reported previously (Fawcett and Camerman, Acta Cryst., 1975, 31, 658–665). (Ala)₄ with AP β‐structure is more stable than AP‐(Ala)₃ due to formation of the stronger hydrogen bonds. The intermolecular structure of (Ala)₄ is also different from polyalanine fiber structure, indicating that the interchain arrangement of AP β‐structure changes with increasing alanine sequencelength. Furthermore the precise ¹H positions, which are usually inaccesible by X‐ray diffraction method, are determined by high resolution ¹H solid state NMR combined with the chemical shift calculations by the gauge‐including projector augmented wave method. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 13–20, 2014.     

Structures and stabilities of ScBn (n = 1–12) clusters: an ab initio investigation

The geometries, stabilities, and electronic properties of ScB_n (n?=?1–12) clusters have been systematically investigated by using density functional theory B3LYP method and coupled–cluster theory CCSD(T) method. It is found that the ground state isomers of ScB_n have planar or quasi–planar structure when n?≤?6, which can be viewed as a B atom of the corresponding B_n+1 cluster is substituted by a Sc atom. From n?≥?7, the ground state isomers favor nest–like structure, in which the Sc atom sits on a nest–like B_n cluster. The calculated second–order differences of energies manifest that the magic numbers of stability are n?=?3, 7, 8, 9 and 11 for the ScB _n clusters. Further analysis indicates that the ScB₇ cluster with C _6v symmetry represents the outstanding stable ScB_n cluster, as confirmed by its electronic structure and molecular orbitals.     

Probing the electronic structures and properties of neutral and charged arsenic sulfides (As n S(−1,0,+1), n = 1–7) using Gaussian-3 theory

The structures and energies of neutral and charged arsenic sulfides As _n S^(?1,0,+1) (n?=?1–7) were systematically investigated using the G3 method. The bonding properties and the stabilities of As _n S and their ions were discussed. The adiabatic electron affinities (AEAs) and adiabatic ionization potentials (AIPs) were presented. The ground-state structures of As _n S can be considered as the lowest-energy structure of neutral As _n+1 by replacing an As atom with a S atom, that is, “substitutional structure”, in which the feature of sulfur bonding is edge-bridging. The ground-state structures of As _n S⁺ tend to be derived from the lowest-energy structure of cation As _n ⁺ by attaching to a S atom, that is, “attaching structure”, in which the sulfur can be three-fold coordinated. There is no rule to be found for the ground-state structure of anion As _n S^?, in which the sulfur can be a terminal atom. There are odd-even alternations in both AEAs and AIPs as a function of size of As _n S. The dissociation energies of S, S^?, and/or S⁺ from neutral As _n S and their ions were calculated to examine their stabilities.     

The βE-Domain of Wheat Ec-1 Metallothionein: A Metal-Binding Domain with a Distinctive Structure

Metallothioneins (MTs) are ubiquitous cysteine-rich proteins with a high affinity for divalent metal ions such as Zn^II, Cu^I, and Cd^II that are involved in metal ion homeostasis and detoxification, as well as protection against reactive oxygen species. Here we show the NMR solution structure of the β_E-domain of the early cysteine-labeled protein (E_c-1) from wheat (β_E-E_c-1), which represents the first three-dimensional structure of a plant MT. The β_E-domain comprises the 51 C-terminal residues of E_c-1 and exhibits a distinctive unprecedented structure with two separate metal-binding centers, a mononuclear Zn^II binding site constituted by two cysteine and two highly conserved histidine residues as found in certain zinc-finger motifs, and a cluster formed by three Zn^II ions coordinated by nine Cys residues that resembles the cluster in the β-domain of vertebrate MTs. Cys-metal ion connectivities were determined by exhaustive structure calculations for all 7560 possible configurations of the three-metal cluster. Backbone dynamics investigated by ¹⁵N relaxation experiments support the results of the structure determination in that β_E-E_c-1 is a rigidly folded polypeptide. To further investigate the influence of metal ion binding on the stability of the structure, we replaced Zn^II with Cd^II ions and examined the effects of metal ion release on incubation with a metal ion chelator.     

Theoretical studies on reaction mechanism of H2 with COS

The reaction mechanisms of H₂ with OCS have been investigated theoretically by using density function theory method. Three possible pathways leading to major products CO and H₂S, as well as two possible pathways leading to by-product CH₄ have been proposed and discussed. For these reaction pathways, the structure parameters, vibrational frequencies and energies for each stationary point have been calculated, and the corresponding reaction mechanism has been given by the potential energy surface, which is drawn according to the relative energies. The calculated results show that the corresponding major products CO and H₂S as well as by-product CH₄ are in agreement with experimental findings, which provided a new illustration and guidance for the reaction of H₂ with OCS.     

Detection of dynamic water molecules in a microcrystalline sample of the SH3 domain of α-spectrin by MAS solid-state NMR

Water molecules are a major determinant of protein stability and are important for understanding protein–protein interactions. We present two experiments which allow to measure first the effective T₂ decay rate of individual amide proton, and second the magnetization build-up rates for a selective transfer from H₂O to H^N using spin diffusion as a mixing element. The experiments are demonstrated for a uniformly ²H, ¹⁵N labeled sample of a microcrystalline SH3 domain in which exchangeable deuterons were back-substituted with protons. In order to evaluate the NMR experimental data, as X-ray structure of the protein was determined using the same crystallization protocol as for the solid-state NMR sample. The NMR experimental data are correlated with the dipolar couplings calculated from H₂O–H^N distances which were extracted from the X-ray structure of the protein. We find that the H^N T₂ decay rates and H₂O–H^N build-up rates are sensitive to distance and dynamics of the detected water molecules with respect to the protein. We show that qualitative information about localization and dynamics of internal water molecules can be obtained in the solid-state by interpretation of the spin dynamics of a reporter amide proton.     

Three-dimensional 1H NMR structure of the nucleocapsid protein NCp10 of Moloney murine leukemia virus

Summary The nucleocapsid protein of Moloney murine leukemia virus (NCp10) is a 56-amino acid protein which contains one zinc finger of the CysX₂CysX₄HisX₄Cys form, a highly conserved motif present in most retroviruses and retroelements. At pH5, NCp10 binds one zinc atom and the complexation induces a folding of the CysX₂CysX₄HisX₄Cys box, similar to that observed for the zinc-binding domains of HIV-1 NC protein. The three-dimensional structure of NCp10 has been determined in aqueous solution by 600 MHz ¹H NMR spectroscopy. The proton resonances could be almost completely assigned by means of phase-sensitive double-quantum-filtered COSY, TOCSY and NOESY techniques. NOESY spectra yielded 597 relevant structural constraints, which were used as input for distance geometry calculations with DIANA. Further refinement was performed by minimization with the program AMBER, which was modified by introducing a zinc force field. The solution structure is characterized by a well-defined central zinc finger (rmsd of 0.747±0.209 Å for backbone atoms and 1.709±0.187 Å when all atoms are considered), surrounded by flexible N- and C-terminal domains. The Tyr²⁸, Trp³⁵, Lys³⁷, Lys⁴¹ and Lys⁴² residues, which are essential for activity, lie on the same face of the zinc finger, forming a bulge structure probably involved in viral RNA binding. The significance of these structural characteristics for the various biological functions of the protein is discussed, taking into account the results obtained with various mutants.     

Hydrothermal synthesis and structural investigation of silver magnesium complex of benzenehexacarboxylic acid (mellitic acid), Ag2Mg2[C6(COO)6] · 8H2O with two-dimensional layered structure

Crystal and molecular structure of silver magnesium mellitate, Ag₂Mg₂[C₆(COO)₆] · 8H₂O, was synthesized hydrothermally and characterized by X-ray structure analysis. The complex crystallizes in the monoclinic system, space group P2/n, with unit cell dimensions of a=7.4347(2), b=9.9858(2), c=14.4248(3) Å, β=99.2429(5)°, V=1055.01(4) ų, and Z=2. The structure was solved and refined to R=0.036 (R_w=0.045) for 1707 independent reflections [I_o>2σ(I_o)]. The Ag cations are coordinated by six carboxylic oxygen atoms of mellitate anions with composition of [C₆(COO)₆]⁶⁻ on the (1 0 1) plane; each mellitate anion linking three neighboring Ag distorted trigonal prisms produces a two-dimensional layered structure parallel to (1 0 1). The Mg cations, which are coordinated by four water molecules and two carboxylic oxygen atoms, are intercalated between the two-dimensional layer stacks. The carboxylate group coordinated to Mg and Ag cations serve as a tridentate ligand in that structure. The number of water molecules incorporated into the mellitate compound is controlled mainly by ionic radii of metal cation in the structure. Furthermore, the ionic radii of metal cations in the mellitate compound play an essential role in arrangement of mellitate anions in the structure, whether as a one-dimensional infinite chain, a two-dimensional layered structure, or a three-dimensional framework structure.     

Partitioning, dynamics, and orientation of lung surfactant peptide KL4 in phospholipid bilayers

Lung surfactant protein B (SP-B) is a lipophilic protein critical to lung function at ambient pressure. KL₄ is a 21-residue peptide which has successfully replaced SP-B in clinical trials of synthetic lung surfactants. CD and FTIR measurements indicate KL₄ is helical in a lipid bilayer environment, but its exact secondary structure and orientation within the bilayer remain controversial. To investigate the partitioning and dynamics of KL₄ in phospholipid bilayers, we introduced CD₃-enriched leucines at four positions along the peptide to serve as probes of side chain dynamics via ²H solid-state NMR. The chosen labels allow distinction between models of helical secondary structure as well as between a transmembrane orientation or partitioning in the plane of the lipid leaflets. Leucine side chains are also sensitive to helix packing interactions in peptides that oligomerize. The partitioning and orientation of KL₄ in DPPC/POPG and POPC/POPG phospholipid bilayers, as inferred from the leucine side chain dynamics, is consistent with monomeric KL₄ lying in the plane of the bilayers and adopting an unusual helical structure which confers amphipathicity and allows partitioning into the lipid hydrophobic interior. At physiologic temperatures, the partitioning depth and dynamics of the peptide are dependent on the degree of saturation present in the lipids. The deeper partitioning of KL₄ relative to antimicrobial amphipathic α-helices leads to negative membrane curvature strain as evidenced by the formation of hexagonal phase structures in a POPE/POPG phospholipid mixture on addition of KL₄. The unusual secondary structure of KL₄ and its ability to differentially partition into lipid lamellae containing varying levels of saturation suggest a mechanism for its role in restoring lung compliance.     

Molecular mechanics calculations of dA12·dT12 and of the curved molecule d(GCTCGAAAAA)4·d(TTTTTCGAGC)4

Using the AMBER software package (Weiner and Kollman 1981) substantially modified for electrostatic contributions, the structural energies of the double-stranded oligonucleotides dA₁₂·dT₁₂ and d(GCTCGAAAAA)₄·d(TTTTTCGAGC)₄ were minimized. Using various starting structures for the molecule dA₁₂·dT₁₂, one final structure is obtained which possesses the experimentally determined properties of poly(dA)·poly(dT). This structure is an A-form-B-form-hybrid structure similar to that of Arnott et al. (1983). The dA-strand is similar to an A-form while the dT-strand is similar to normal B-form. This structure and separately optimized B-form sequence stretches were used to construct the double-stranded fragment d(GCTCGAAAAA)₄ which again was optimized. This sequence, when imbedded in a DNA fragment as contiguous repeats, shows a gel migration anomaly which has been interpreted as stable curvature of the DNA (Diekmann 1986). The calculated structure of this sequence indeed has a curved helix axis and is discussed as a model for curved DNA. A theoretical formalism is presented which allows one to calculate the structural parameters of any nucleic acid double helix in two different geometrical representations. This formalism is used to determine the parameters of the base-pair orientations of the curved structure in terms of wedge as well as cylindrical parameters. In the structural model presented here, the curvature of the helix axis results from an alternation of two different DNA structures in which the base-pairs possess different angles with the helix axis (cylinder tilt). Resulting from geometric restraints, a negative cylinder tilt angle correlates strongly with the closing of the minor groove (wedge roll). The blocks with different structure are not exactly coincident with the dA₅-blocks and the B-DNA stretches. Within the dA₅ block, base-pair tilt and wedge roll adopt large values which proceed into the 3 flanking B-DNA sequence by about one base-pair. These properties of the structure calculated here are discussed in terms of different models explaining DNA curvature.     

Conservative secondary structure motifs already present in early-stage folding (in silico) as found in serpines family

The well-known procedure implemented in ClustalW oriented on the sequence comparison was applied to structure comparison. The consensus sequence as well as consensus structure has been defined for proteins belonging to serpine family. The structure of early stage intermediate was the object for similarity search. The high values of W_sequence appeared to be accordant with high values of W_structure making possible structure comparison using common criteria for sequence and structure comparison.Since the early stage structural form has been created according to limited conformational sub-space which does not include the β-structure (this structure is mediated by C7eq structural form), is particularly important to see, that the C7eq structural form may be treated as the seed for β-structure present in the final native structure of protein.The applicability of ClustalW procedure to structure comparison makes these two comparisons unified.     

Synthesis and characterization of [Fe2OX6]2tau; (X = Cl,Br, I) complexes. Crystal and molecular structure of (BzPh3P)2 [Fe2OCl6]

Improved and simple methods for the preparation of [Fe₂OX₆]^2tau; (X = Cl Br I) complexes are described. The complexes were obtained in high and purified yields as the BzPh₃P⁺ or R₄N⁺ salts. An X-ray crystallographic study of (BzPh₃P)₂ [Fe₂OCl₆] revealed a structure for the complex anion in which the two iron atoms are linked by gdot; -oxo bridge and the terminal coordination sites are occupied by the chloride ligands. Mcolon; ssbauer and infrared spectra for the complexes are reported.     

Biogenic hierarchical TiO₂/SiO₂ derived from rice husk and enhanced photocatalytic properties for dye degradation

Background

Rice husk, an agricultural bioresource, is utilized as a non-metallic bio-precursor to synthesize biogenic hierarchical TiO₂/SiO₂ (BH-TiO₂/SiO₂) and the products are applied to dye degradation.

Methodology/Principal Findings

The as-prepared BH-TiO₂/SiO₂ samples are characterized by X-ray diffraction(XRD), X-ray photoelectron spectroscopy(XPS), nitrogen-adsorption measurement, UV-vis spectroscopy and electronic paramagnetic resonance (EPR). The results show that BH-TiO₂/SiO₂ possesses both anatase and rutile phases with amorphous SiO₂ as background, which contains mesopore structure, and nitrogen derived from original rice husk is self-doped into the lattice. Besides, the light-harvesting within the visible-light range of BH-TiO₂/SiO₂ has been enhanced. Moreover, the catalytic activity of BH-TiO₂/SiO₂ has been proven by EPR, and both the photocatalytic activity and stability of BH-TiO₂/SiO₂ are improved as well, which has been illustrated by cycled degradation of methylene blue dye under irradiation.

Conclusions/Significance

This work provides a good way to combine natural hierarchical porous structure with synthetic material chemistry based on available biomass in the vast natural environment for the sustainable development of human society, and extends potentials of biomass in applications such as photocatalysts, sunlight splitting water and so forth.     

Monodisperse and LPS-free Aggregatibacter actinomycetemcomitans leukotoxin: Interactions with human β2 integrins and erythrocytes

Aggregatibacter actinomycetemcomitans is a gram-negative, facultatively anaerobic cocco-bacillus and a frequent member of the human oral flora. It produces a leukotoxin, LtxA, belonging to the repeats-in-toxin (RTX) family of bacterial cytotoxins. LtxA efficiently kills neutrophils and mononuclear phagocytes. The known receptor for LtxA on leukocytes is integrin α_Lβ₂ (LFA-1 or CD11a/CD18). However, the molecular mechanisms involved in LtxA-mediated cytotoxicity are poorly understood, partly because LtxA has proven difficult to prepare for experiments as free of contaminants and with its native structure. Here, we describe a protocol for the purification of LtxA from bacterial culture supernatant, which does not involve denaturing procedures. The purified LtxA was monodisperse, well folded as judged by the combined use of synchrotron radiation circular dichroism spectroscopy (SRCD) and in silico prediction of the secondary structure content, and free of bacterial lipopolysaccharide. The analysis by SRCD and similarity to a lipase from Pseudomonas with a known three dimensional structure supports the presence of a so-called beta-ladder domain in the C-terminal part of LtxA. LtxA rapidly killed K562 target cells transfected to express β₂ integrin. Cells expressing α_Mβ₂ (CD11b/CD18) or α_Xβ₂ (CD11c/CD18) were killed as efficiently as cells expressing α_Lβ₂. Erythrocytes, which do not express β₂ integrins, were lysed more slowly. In ligand blotting experiments, LtxA bound only to the β₂ chain (CD18). These data support a previous suggestion that CD18 harbors the major binding site for LtxA as well as identifies integrins α_Mβ₂ and α_Xβ₂ as novel receptors for LtxA.     

Spectroscopic and magnetic properties of transition metal(II) triflouromethanesulfonate compounds with bridging asymmetric 3,4-dialkyl substituted 1,2,4-triazole ligands

The spectroscopic and magnetic properties are described of coordination compounds with asymmetric 3,4-diakyl substituted 1,2,4-triazoles. The ligands 3-methyl-4-ethyl-1,2,4-traizole and 3-methyl-4-t-butyl-1,2,4-triazole have been investigated. Using M(CF₃SO₃)₂ (M = Mn, Co, Ni, Cu, Zn) compounds have been obtained with a linear trinuclear structure, in which the metal ions are linked to each other by two pairs of three bridging triazoles. The coordination sphere around the terminal ligands is completed by monodentate ligands and/or water molecules. The structure has been confirmed by an X-ray structure determination of [Co₃(metz)₆(H₂O)₆][Co₃ (metz)₈(H₂O)₄](CF₃SO₃)₁₂(H₂O)₈. this compound crystallizes in the space group P with lattice constants a = 13.793(4), b = 14.401(3), c = 23.258(4) Å, α = 80.58(2), β = 83.23(2) and γ = 64.33(2)°. The unit cell contains two independent trinuclear clusters of different composition. These two clusters have the same overall structure. Differences are related to the presence of monodentately coordinating ligands as well as to the position of the C3-methyl substituent. The complete refinement of this structure was obstructed by disorder problems in the anions and the non-coordinating water molecules. The magnetic susceptibilities of the compounds have been recorded and could be fitted to theoretical expressions for linear trimers. The compound [Ni₂- (mtbtz)₅(H₂O)₄](CF₃SO₃)₄(H₂O)₄ appears to be a dimeric species as concluded from its magnetic behaviour.     

Structure and thermal behavior of the layered zincophosphate [NH3-CH2-CH(NH3)-CH3](ZnPO4)2

A zinc phosphate with the composition [NH₃-CH₂-CH(NH₃)-CH₃](ZnPO₄)₂, containing the doubly protonated 1,2-diaminopropane (abbr. HDAP), was synthesized by hydrothermal crystallization of zinc nitrate, phosphoric acid, 1,2-diaminopropane and trimethylenedipiperidine. The single crystal analysis shows a layered inorganic-organic structure built up of a sandwich-like motif of alternating inorganic layers and HDAP cations (trimethylenedipiperidine not being a constituent of the structure). The HDAP species are found to be disordered and serve as bridges between two adjacent inorganic layers which are separated by 3.95 Å. The bridging interaction occurs via a hydrogen-bonding network. The inorganic layer features a pattern of four-membered rings involving two ZnO₄ and two PO₄ tetrahedra connected by sharing O atoms. Thermal analysis shows that the compound is stable up to 370 °C and that the thermal decomposition of HDAP occurs in two steps between 370 and 460 °C, causing the collapse of the structure. The first decomposition step corresponds to ammonia removal which proceeds with a high activation energy (E_a = 282 kJ mol⁻¹). The high E_a value is mainly attributed to strong electrostatic interactions between organic cations and anionic inorganic layers, the disruption of the interactions being the main reason for structural collapse after the HDAP removal.     

The Configurational Space of Rocksalt‐Type Oxides for High‐Capacity Lithium Battery Electrodes

A unifying theory is presented to explain the lithium exchange capacity of rocksalt‐like structures with any degree of cation ordering, and how lithium percolation properties can be used as a guideline for the development of novel high‐capacity electrode materials is demonstrated. The lithium percolation properties of the three most common lithium metal oxide phases, the layered α‐NaFeO₂ structure, the spinel‐like LT‐LiCoO₂ structure, and the γ‐LiFeO₂ structure, are demonstrated and a strong dependence of the percolation thresholds on the cation ordering and the lithium content is observed. The poor performance of γ‐LiFeO₂‐type structures is explained by their lack of percolation of good Li migration channels. The spinel‐like structure exhibits excellent percolation properties that are robust with respect to off‐stoichiometry and some amount of cation disorder. The layered structure is unique, as it possesses two different types of lithium diffusion channels, one of which is, however, strongly dependent on the lattice parameters, and therefore very sensitive to disorder. In general it is found that a critical Li‐excess concentration exists at which Li percolation occurs, although the amount of Li excess needed depends on the partial cation ordering. In fully cation‐disordered materials, macroscopic lithium diffusion is enabled by ≈10% excess lithium.     

Crystal structure and exchange pathways of the complex l-(trypthophyl-glycinato)copper(II)

The crystal structure and the EPR characterization of the compound Cu [C₁₃H₁₃N₃O₃] is reported. It crystallizes in the P2₁2₁2₁ space group, with a=8.2829(5), b=9.347(2), c=16.499(2) Å and Z=4. The copper ion is in a distorted square planar coordination, bonded to two nitrogen and one oxygen atoms from one dipeptide and to an oxygen atom from a symmetry-related molecule. Thus, neighbor copper atoms at 5.14 Å are connected by equatorial syn–anti carboxylate bridges giving rise to a chain structure along the b-axis. The chains are connected via hydrogen bonds and cation–π interactions, the latter being provided by the ‘sandwich’ structure involving each copper atom and two tryptophan residues from neighbor molecules. The EPR spectra of polycrystalline sample imply an essentially d_x2−y2 ground state orbital for the Cu(II) ions. The g-values reflect a slightly distorted axial symmetry around the Cu(II) ions as expected from the structural results. No hyperfine interaction is observed, which is indicative of the presence of exchange interactions between the copper atoms as suggested by the X-ray results as well.     

Solution structure of a DNA quadruplex containing ALS and FTD related GGGGCC repeat stabilized by 8-bromodeoxyguanosine substitution

A prolonged expansion of GGGGCC repeat within non-coding region of C9orf72 gene has been identified as the most common cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), which are devastating neurodegenerative disorders. Formation of unusual secondary structures within expanded GGGGCC repeat, including DNA and RNA G-quadruplexes and R-loops was proposed to drive ALS and FTD pathogenesis. Initial NMR investigation on DNA oligonucleotides with four repeat units as the shortest model with the ability to form an unimolecular G-quadruplex indicated their folding into multiple G-quadruplex structures in the presence of K⁺ ions. Single dG to 8Br-dG substitution at position 21 in oligonucleotide d[(G₄C₂)₃G₄] and careful optimization of folding conditions enabled formation of mostly a single G-quadruplex species, which enabled determination of a high-resolution structure with NMR. G-quadruplex structure adopted by d[(G₄C₂)₃GG^BrGG] is composed of four G-quartets, which are connected by three edgewise C-C loops. All four strands adopt antiparallel orientation to one another and have alternating syn-anti progression of glycosidic conformation of guanine residues. One of the cytosines in every loop is stacked upon the G-quartet contributing to a very compact and stable structure.