Unconventional Interactions Research Articles

The expanding universe of ribonucleoproteins: of novel RNA-binding proteins and unconventional interactions.

Post-transcriptional regulation of gene expression plays a critical role in almost all cellular processes. Regulation occurs mostly by RNA-binding proteins (RBPs) that recognise RNA elements and form ribonucleoproteins (RNPs) to control RNA metabolism from synthesis to decay. Recently, the repertoire of RBPs was significantly expanded owing to methodological advances such as RNA interactome capture. The newly identified RNA binders are involved in diverse biological processes and belong to a broad spectrum of protein families, many of them exhibiting enzymatic activities. This suggests the existence of an extensive crosstalk between RNA biology and other, in principle unrelated, cell functions such as intermediary metabolism. Unexpectedly, hundreds of new RBPs do not contain identifiable RNA-binding domains (RBDs), raising the question of how they interact with RNA. Despite the many functions that have been attributed to RNA, our understanding of RNPs is still mostly governed by a rather protein-centric view, leading to the idea that proteins have evolved to bind to and regulate RNA and not vice versa. However, RNPs formed by an RNA-driven interaction mechanism (RNA-determined RNPs) are abundant and offer an alternative explanation for the surprising lack of classical RBDs in many RNA-interacting proteins. Moreover, RNAs can act as scaffolds to orchestrate and organise protein networks and directly control their activity, suggesting that nucleic acids might play an important regulatory role in many cellular processes, including metabolism.

Submonolayer Ag films on Fe(100): A first-principles analysis of energetics controlling adlayer thermodynamics and kinetics

Epitaxial growth of Ag on Fe(100) and postdeposition relaxation have been studied in several experiments. We provide a first-principles density functional theory analysis of key adatom interaction energies and diffusion barriers controlling growth and relaxation kinetics for the submonolayer regime, as these have not been assessed previously. A cluster expansion approach is used to obtain an extensive set of conventional lateral interactions between adatoms on fourfold hollow adsorption sites. We find robust oscillatory decay of pair interactions with increasing separation, and of trio interactions with increasing perimeter length. First- and second-nearest-neighbor pair interactions, as well as compact linear and bent trio interactions, dominate. The adatom terrace diffusion barrier is estimated to be ${E}_{\mathrm{d}}\ensuremath{\approx}0.39$ eV. We also provide a limited analysis of unconventional interactions for which one adatom is at the bridge-site transition state for hopping and one or more others are at fourfold hollow sites. Energy barriers for diffusion along island edges can be determined with the aid of both conventional and unconventional interactions.

IgG Conformer's Binding to Amyloidogenic Aggregates.

Amyloid-reactive IgGs isolated from pooled blood of normal individuals (pAbs) have demonstrated clinical utility for amyloid diseases by in vivo targeting and clearing amyloidogenic proteins and peptides. We now report the following three novel findings on pAb conformer's binding to amyloidogenic aggregates: 1) pAb aggregates have greater activity than monomers (HMW species > dimers > monomers), 2) pAbs interactions with amyloidogenic aggregates at least partially involves unconventional (non-CDR) interactions of F(ab) regions, and 3) pAb's activity can be easily modulated by trace aggregates generated during sample processing. Specifically, we show that HMW aggregates and dimeric pAbs present in commercial preparations of pAbs, intravenous immunoglobulin (IVIg), had up to ~200- and ~7-fold stronger binding to aggregates of Aβ and transthyretin (TTR) than the monomeric antibody. Notably, HMW aggregates were primarily responsible for the enhanced anti-amyloid activities of Aβ- and Cibacron blue-isolated IVIg IgGs. Human pAb conformer's binding to amyloidogenic aggregates was retained in normal human sera, and mimicked by murine pAbs isolated from normal pooled plasmas. An unconventional (non-CDR) component to pAb's activity was indicated from control human mAbs, generated against non-amyloid targets, binding to aggregated Aβ and TTR. Similar to pAbs, HMW and dimeric mAb conformers bound stronger than their monomeric forms to amyloidogenic aggregates. However, mAbs had lower maximum binding signals, indicating that pAbs were required to saturate a diverse collection of binding sites. Taken together, our findings strongly support further investigations on the physiological function and clinical utility of the inherent anti-amyloid activities of monomeric but not aggregated IgGs.

Importance of polarization assisted/resonance assisted hydrogen bonding interactions and unconventional interactions in crystal formations of five new complexes bearing chelidamic acid through a proton transfer mechanism

Five new coordination compounds derived from chelidamic acid and amines have been synthesized and X-ray characterized. The noncovalent interactions that govern the crystal packing have been rationalized by means of DFT calculations.

Nanoscale Fluid Mechanics and Energy Conversion

Under nanoconfinement, fluid molecules and ions exhibit radically different configurations, properties, and energetics from those of their bulk counterparts. These unique characteristics of nanoconfined fluids, along with the unconventional interactions with solids at the nanoscale, have provided many opportunities for engineering innovation. With properly designed nanoconfinement, several nanofluidic systems have been devised in our group in the past several years to achieve energy conversion functions with high efficiencies. This review is dedicated to elucidating the unique characteristics of nanofluidics, introducing several novel nanofluidic systems combining nanoporous materials with functional fluids, and to unveiling their working mechanisms. In all these systems, the ultra-large surface area available in nanoporous materials provides an ideal platform for seamlessly interfacing with nanoconfined fluids, and efficiently converting energy between the mechanical, thermal, and electrical forms. These systems have been demonstrated to have great potentials for applications including energy dissipation/absorption, energy trapping, actuation, and energy harvesting. Their efficiencies can be further enhanced by designing efforts based upon improved understanding of nanofluidics, which represents an important addition to classical fluid mechanics. Through the few systems exemplified in this review, the emerging research field of nanoscale fluid mechanics may promote more exciting nanofluidic phenomena and mechanisms, with increasing applications by encompassing aspects of mechanics, materials, physics, chemistry, biology, etc.

Unconventional Interaction Forces in Protein and Protein-ligand Systems and their Impacts to Drug Design

In drug design and enzyme engineering, the information of interactions between receptors and ligands is crucially important. In many cases, the protein structures and drug-target complex structures are determined by a delicate balance of several weak molecular interaction types. Among these interaction forces several unconventional interactions play important roles, however, less familiar for researchers. The cation-π interaction is a unique noncovalent interaction only acting between aromatic amino acids and organic cations (protonated amino acids) and inorganic cations (proton and metallic). This article reports new study results in the interaction strength, the behaviors and the structural characters of cation-π interactions between aromatic amino acids (Phe, Tyr, and Trp) and organic and inorganic cations (Lys(+), Arg(+), H(+), H3O(+), Li(+), Na(+), K(+), Ca(2+), and Zn(2+)) in gas phase and in solutions (water, acetonitrile, and cyclohexane). Systematical research revealed that the cation-π interactions are point-to-plane (aromatic group) interactions, distance and orientationdependent, and the interaction energies change in a broad range. In gas phase the cation-π interaction energies between aromatic amino acids (Phe, Tyr, and Trp) and metallic cations (Li(+), Na(+), K(+), Ca(2+), and Zn(2+)) are in the range -12 to -160 kcal/mol, and the interaction energies of protonated amino acids (Arg(+) and Lys(+)) are in the range from -9 to -18 kcal/mol. In solutions the cation-π energies decrease with the dielectric constant ε of solvents. However, in aqueous solution the cation-π energies of H3O(+) and protonated amino acids are less affected by solvation effects. The applications of unconventional interaction forces in drug design and in protein engineering are introduced.

Structure diversities of ten entangled coordination polymers assembled from reactions of Co(ii) or Ni(ii) salts with 5-(pyridin-4-yl)isophthalic acid in the absence or presence of auxiliary N-donor ligands

The self-assembly of transition metal salts with 5-(pyridin-4-yl)isophthalate ligand and N-donor ligands affords a series of entangled coordination frameworks based on different metal cluster cores, namely, [Co(L)(H2O)2]·H2O (1), [Ni(L)(H2O)]·H2O (2), [Co4(L)2(SO4)(H2O)5(OH)2]·15.5H2O (3), [Co2(L)2(2,2′-bpy)2]2·H2O (4), [Co2(L)2(phen)2] (5), [Ni(L)(en)(H2O)2]2·2H2O (6), [Ni(L)(dap)]·3H2O (7), [Co(L)(bph)(H2O)2]·H2O (8), [Co(L)(bbm)0.5]·4H2O (9), [Co(L)(bimb)0.5]4·13H2O (10) [L = 5-(pyridin-4-yl)isophthalic acid; 2,2′-bpy = 2,2′-bipyridine; phen = 1,10-phenanthroline; en = ethane-1,2-diamine; dap = propane-1,2-diamine; bph = (1E,2E)-1,2-bis(pyridin-4-ylmethylene)hydrazine; bbm = 1,4-di(1H-imidazol-1-yl)benzene; bimb = 4,4′-di(1H-imidazol-1-yl)-1,1′-biphenyl]. Their structures were characterized by elemental analysis, IR spectra and TG analyses and further determined by single-crystal X-ray diffraction analysis. The auxiliary ligands play important roles in the formation of various entangled networks. 1–2 show a 2D layer network and its layers are held together by π–π stacking interactions and other unconventional interactions. Complex 3 displays an unusual Co4 cluster core which is linked by η1,η1,η1,μ3-SO42− and μ3-OH ions. Upon adding chelating ligands, 4–5 show 2D → 2D entangled polymers with polycatenated and polyrotaxane features based on 4,4-sql layers. 6–7 still exhibit layer strutures, although they have chelating ligands, which may be due to the absence of π–π stacking interactions. 8–10 present pillar-layer structures which are supported by different lengths of pillars. Three-fold interpenetration with polycatenated features are observed in 8–9. 10 exhibits a 4-fold interpenetrating 3D structure with large channel running through its net.

Direct observation of hydrogen atom dynamics and interactions by ultrahigh resolution neutron protein crystallography

The 1.1 Å, ultrahigh resolution neutron structure of hydrogen/deuterium (H/D) exchanged crambin is reported. Two hundred ninety-nine out of 315, or 94.9%, of the hydrogen atom positions in the protein have been experimentally derived and resolved through nuclear density maps. A number of unconventional interactions are clearly defined, including a potential O─H…π interaction between a water molecule and the aromatic ring of residue Y44, as well as a number of potential C─H…O hydrogen bonds. Hydrogen bonding networks that are ambiguous in the 0.85 Å ultrahigh resolution X-ray structure can be resolved by accurate orientation of water molecules. Furthermore, the high resolution of the reported structure has allowed for the anisotropic description of 36 deuterium atoms in the protein. The visibility of hydrogen and deuterium atoms in the nuclear density maps is discussed in relation to the resolution of the neutron data.

Does the oestrogen receptor encourage oestrogenicity in environmental pollutants? The case of 4-nonylphenol

A computer-aided docking study was conducted to explore in detail the binding interactions between the structurally unlikely environmental oestrogen 4-nonylphenol (4NP) and three of its metabolites with the human oestrogen receptor alpha (hERα). Docking was done within the Schrodinger Suite 2008 using both a conventional rigid receptor with flexible ligand and the induced-fit docking protocol. Induced-fit docking allows side-chain and backbone movement in the receptor to accommodate the ligand. This study has revealed unconventional interactions between the ligands and the hERα binding pocket that could explain the observed oestrogen-like behaviour of 4NP and suggests some of the metabolites of 4NP may also be oestrogenic.

Many Paths to the Same End: Histone Transcripts Recruit Canonical Initiation Factors through Unconventional Interactions

In this issue of Molecular Cell, Eriani and colleagues show that histone transcripts recruit translation initiation factors through an alternate pathway reminiscent of viral translation, suggesting a broader role for noncanonical translation initiation on cellular transcripts (Martin etal., 2011).

Supramolecular frameworks constructed from angular-shaped sulfonyldipyrazine and various silver(i) salts

The new angular-shaped multidentate ligand sulfonyldipyrazine (abbreviated as L) is capable of forming diverse coordination polymeric motifs with various silver(I) salts, such as the infinite chain structure of [Ag3(L)(NO2)3]∞, (4,4) net of isomorphous {[Ag(L)](ClO4)}∞ and [Ag(L)(NO3)]∞, and three-dimensional frameworks of [Ag(L)(CF3SO3)]∞ and [Ag(L)(CF3CO2)]∞. In these complexes, sulfonyldipyrazine exhibits flexible ligation modes with its pyrazinyl N atoms bound to adjacent metal centers. The electron-deficient pyrazinyl rings and the sulfonyl group at the bend provide additional bonding sites for unconventional interactions, such as anion–π(pyrazinyl) and SO⋯π(pyrazinyl) interactions, in the supramolecular assembly of these complexes.

Oriented Immobilization of Prion Protein Demonstrated via Precise Interfacial Nanostructure Measurements

Nanopatterning of biomolecules on functionalized surfaces offers an excellent route for ultrasensitive protein immobilization, for interaction measurements, and for the fabrication of devices such as protein nanoarrays. An improved understanding of the physics and chemistry underlying the device properties and the recognition process is necessary for performance optimization. This is especially important for the recognition and immobilization of intrinsically disordered proteins (IDPs), like the prion protein (PrP), a partial IDP, whose folding and stability may be influenced by local environment and confinement. Atomic force microscopy allows for both highly controllable nanolithography and for sensitive and accurate direct detection, via precise topographic measurements on ultraflat surfaces, of protein interactions in a liquid environment, thus different environmental parameters affecting the biorecognition phenomenon can be investigated in situ. Using nanografting, a tip-induced lithographic technique, and an affinity immobilization strategy based on two different histidine tagged antibodies, with high nM affinity for two different regions of PrP, we successfully demonstrated the immobilization of recombinant mouse PrP onto nanostructured surfaces, in two different orientations. Clear discrimination of the two molecular orientations was shown by differential height (i.e., topographic) measurements, allowing for the estimation of binding parameters and the full characterization of the nanoscale biorecognition process. Our work opens the way to several high sensitivity diagnostic applications and, by controlling PrP orientation, allows for the investigation of unconventional interactions with partially folded proteins, and may serve as a platform for protein misfolding and refolding studies on PrP and other thermodynamically unstable, fibril forming, proteins.

Synthesis, structure, and estrogenic activity of 4-amino-3-(2-methylbenzyl)coumarins on human breast carcinoma cells

A number of coumarins exhibit interesting pharmacological activities and are therefore of therapeutic use. We report here the synthesis and the structural analysis of new N-substituted 4-amino-3-(2-methylbenzyl)coumarins (compounds 8a– 8e) that present structural analogies with estrothiazine and 11- or 7-substituted 17β-estradiol. These derivatives were tested with respect to estrogenic activity on the estrogen receptor positive (ER+) human MCF-7 breast cancer cell line. Two of the reported compounds ( 8a and 8b) stimulated specifically the proliferation of MCF-7 cells, but not that of estrogen receptor negative (ER−) human MDA-MB-231 breast cancer cells, suggesting that their mitogenic activity is mediated by ER. Accordingly, the stimulating effect of 8a and 8b was suppressed by the pure antiestrogen fulvestrant. Besides, 8a and 8b induced ER down-regulation similar to that produced by classical ER agonists or pure antagonists. The effects of the compounds under study on ER-mediated transcription were assessed on (ER+) MVLN cells, that is, MCF-7 cells stably transfected with a pVit-tk-Luc reporter plasmid. Derivatives 8a and 8b, and surprisingly compound 8c, enhanced ER-mediated gene transactivation in that model. Finally, no coumarin was able to compete with tritiated 17β-estradiol ([ 3H]E 2) for ER binding, suggesting unconventional interactions with the receptor, such as interactions with the second binding pocket or with the coactivator-binding region. To conclude, observations performed in this study on compound 8c reveal that estrogenic activity can be dissociated from enhancement of cell proliferation. Furthermore, ERE-driven transactivation of transcription seems to be a condition necessary, but not sufficient, for estrogen-induced stimulation of cell growth.

Molecular Basis of TCR Selectivity, Cross-Reactivity, and Allelic Discrimination by a Bacterial Superantigen: Integrative Functional and Energetic Mapping of the SpeC-Vβ2.1 Molecular Interface

Superantigens activate large fractions of T cells through unconventional interactions with both TCR beta-chain V domains (Vbetas) and MHC class II molecules. The bacterial superantigen streptococcal pyrogenic exotoxin C (SpeC) primarily stimulates human Vbeta2(+) T cells. Herein, we have analyzed the SpeC-Vbeta2.1 interaction by mutating all SpeC residues that make contact with Vbeta2.1 and have determined the energetic and functional consequences of these mutations. Our comprehensive approach, including mutagenesis, functional readouts from both bulk T cell populations, and an engineered Vbeta2.1(+) Jurkat T cell, as well as surface plasmon resonance binding analysis, has defined the SpeC "functional epitope" for TCR engagement. Although only two SpeC residues (Tyr(15) and Arg(181)) are critical for activation of virtually all human CD3(+) T cells, a larger cluster of four hot spot residues are required for interaction with Vbeta2.1. Three of these residues (Tyr(15), Phe(75), and Arg(181)) concentrate their binding energy on the CDR2 loop residue Ser(52a), a noncanonical residue insertion found only in Vbeta2 and Vbeta4 chains. Plasticity of this loop is important for recognition by SpeC. Although SpeC interacts with the Vbeta2.1 hypervariable CDR3 loop, our data indicate these contacts have little to no influence on the functional interaction with Vbeta2.1. These studies also provide a molecular basis for selectivity and cross-reactivity of SpeC-TCR recognition and reveal a degree of fine specificity in these interactions, whereby certain SpeC mutants are capable of distinguishing between different alleles of the same Vbeta domain subfamily.

Mitogen-activated protein kinases interacting kinases are autoinhibited by a reprogrammed activation segment

Autoinhibition is a recurring mode of protein kinase regulation and can be based on diverse molecular mechanisms. Here, we show by crystal structure analysis, nuclear magnetic resonance (NMR)-based nucleotide affinity studies and rational mutagenesis that nonphosphorylated mitogen-activated protein (MAP) kinases interacting kinase (Mnk) 1 is autoinhibited by conversion of the activation segment into an autoinhibitory module. In a Mnk1 crystal structure, the activation segment is repositioned via a Mnk-specific sequence insertion at the N-terminal lobe with the following consequences: (i) the peptide substrate binding site is deconstructed, (ii) the interlobal cleft is narrowed, (iii) an essential Lys-Glu pair is disrupted and (iv) the magnesium-binding loop is locked into an ATP-competitive conformation. Consistently, deletion of the Mnk-specific insertion or removal of a conserved phenylalanine side chain, which induces a blockade of the ATP pocket, increase the ATP affinity of Mnk1. Structural rearrangements required for the activation of Mnks are apparent from the cocrystal structure of a Mnk2 D228G -staurosporine complex and can be modeled on the basis of crystal packing interactions. Our data suggest a novel regulatory mechanism specific for the Mnk subfamily.

Study on β-sheet packing, stabilized by aromatic interactions in a tri-peptide crystal

Abstract The role of unconventional aromatic interactions in the β-sheet packing, inside the crystal, has been highlighted. Crystal structure of terminally blocked tri-peptide, Z-L-Ala-L-Ala-L-Leu-pNA has been determined. There are four different conformers of tri-peptide inside the unit-cell, in space group P1. All the four independent molecules are described by semi-extended backbone conformations which form an anti-parallel β-sheet. The inter β-sheet packing is predominantly stabilized by C—H…π and π…π interactions. In π…π interactions, the center-to-center distance between aromatic rings varies from 3.8 Å to 4.6 Å while the closest distance of approach ranges from 3.4 Å to 3.8 Å. The associations of aromatic-aromatic rings are described by either face-to-face or inclined arrangements.

Unconventional interactions between water and heterocyclic nitrogens in protein structures.

We report an unusual interaction in which a water molecule approaches the heterocyclic nitrogen of tryptophan and histidine along an axis that is roughly perpendicular to the aromatic plane of the side chain. The interaction is distinct from the well-known conventional aromatic hydrogen-bond, and it occurs at roughly the same frequency in protein structures. Calculations indicate that the water-indole interaction is favorable energetically, and we find several cases in which such contacts are conserved among structural orthologs. The indole-water interaction links side chains and peptide backbone in turn regions, connects the side chains in beta-sheets, and bridges secondary elements from different domains. We suggest that the water-indole interaction can be indirectly responsible for the quenching of tryptophan fluorescence that is observed in the folding of homeodomains and, possibly, many other proteins. We also observe a similar interaction between water and the imidazole nitrogens of the histidine side chain. Taken together, these observations suggest that the unconventional water-indole and water-imidazole interactions provide a small but favorable contribution to protein structures.

CD5 and other superantigens as 'ticklers' of the B-cell receptor.

Abstract The exact roles of endogenous and exogenous ligands as stimuli for B-cell survival through ‘tickling' the B-cell receptor are unclear. This article considers the contribution of unconventional interactions with the variable heavy chain by CD5 and other superantigens.

CD5 and other superantigens as ‘ticklers’ of the B-cell receptor

The exact roles of endogenous and exogenous ligands as stimuli for B-cell survival through ‘tickling’ the B-cell receptor are unclear. This article considers the contribution of unconventional interactions with the variable heavy chain by CD5 and other superantigens.

All human tRNATyr genes contain introns as a prerequisite for pseudouridine biosynthesis in the anticodon.

Two synthetic oligonucleotides, one specific for the 5' exon, the other spanning the splice junction, were used to show that (a) the human haploid genome contains at least 12 independent gene loci for tRNATyr, and (b) that all of them carry an intron. From one of the cloned human tRNATyr genes (pHtT1) the 20 bp intron was deleted to generate pHtT1 delta. Homologous in vitro transcription, fingerprint analyses of the products and elucidation of their nucleoside composition revealed that the pseudouridine (psi 35) in the center of the anticodon of tRNATyr was synthesized in the intron-containing precursor whereas this U to psi modification did not take place in precursors or mature tRNATyr derived from pHtT1 delta. On the basis of these results and of studies from other laboratories we suggest that the evolutionary pressure for maintaining introns in eukaryotic tRNAsTyr is this strict intron-requirement for psi 35 synthesis. Taking into account that all eukaryotic cytoplasmic tRNAsTyr contain a psi 35 we discuss here a special need for this modified nucleoside in stabilizing codon-anticodon interactions involving (a) classical base pairing upon translation of tyrosine codons and (b) unconventional interactions during UAG amber codon suppression by tRNATyrG psi A in eukaryotic cells.