Fold Structures Research Articles

Single turnover transient state kinetics reveals processive protein unfolding catalyzed by Escherichia coli ClpB.

Escherichia coli ClpB and Saccharomyces cerevisiae Hsp104 are AAA+ motor proteins essential for proteome maintenance and thermal tolerance. ClpB and Hsp104 have been proposed to extract a polypeptide from an aggregate and processively translocate the chain through the axial channel of its hexameric ring structure. However, the mechanism of translocation and if this reaction is processive remains disputed. We reported that Hsp104 and ClpB are non-processive on unfolded model substrates. Others have reported that ClpB is able to processively translocate a mechanically unfolded polypeptide chain at rates over 240 amino acids (aa) per second. Here, we report the development of a single turnover stopped-flow fluorescence strategy that reports on processive protein unfolding catalyzed by ClpB. We show that when translocation catalyzed by ClpB is challenged by stably folded protein structure, the motor enzymatically unfolds the substrate at a rate of ~0.9 aa s-1 with a kinetic step-size of ~60 amino acids at sub-saturating [ATP]. We reconcile the apparent controversy by defining enzyme catalyzed protein unfolding and translocation as two distinct reactions with different mechanisms of action. We propose a model where slow unfolding followed by fast translocation represents an important mechanistic feature that allows the motor to rapidly translocate up to the next folded region or rapidly dissociate if no additional fold is encountered.

Tunable wave coupling in periodically rotated Miura-ori tubes.

Origami folding structures are vital in shaping programmable mechanical material properties. Of particular note, tunable dynamical properties of elastic wave propagation in origami structures have been reported. Despite the promising features of origami metamaterials, the influence of the kinematics of tessellated origami structures on elastic wave propagation remain unexplored. This study proposes elastic metamaterials using connected Miura-ori tubes, the kinematics of which are coupled by folding and unfolding motions in a tubular axis; achieved by periodically connecting non-rotated and rotated Miura-ori tubes. The kinematics generate wave modes with localized deformations within the unit cell of the metamaterials, affecting the global elastic deformation of Miura-ori tubes via the coupling of wave modes. Dispersion analysis, using the generalized Bloch wave framework based on bar-and-hinge models, verifies the influence of kinematics in the connected tubes on elastic wave propagation. Furthermore, folding the connected tubes changes the coupling strength of wave modes between the kinematics and global elastic deformation of the tubes by breaking the ideal kinematics. The coupling of wave modescontributes to the formation of the band gaps and their tunability. These findings enable adaptive and in situ tunability of band structures to prohibit elastic waves in the desired frequency ranges.This article is part of the theme issue 'Origami/Kirigami-inspired structures: from fundamentals to applications'.

Self-assembly and condensation of intermolecular poly(UG) RNA quadruplexes.

Poly(UG) or 'pUG' dinucleotide repeats are highly abundant sequences in eukaryotic RNAs. In Caenorhabditis elegans, pUGs are added to RNA 3' ends to direct gene silencing within Mutator foci, a germ granule condensate. Here, we show that pUG RNAs efficiently self-assemble into gel condensates through quadruplex (G4) interactions. Short pUG sequences form right-handed intermolecular G4s (pUG G4s), while longer pUGs form left-handed intramolecular G4s (pUG folds). We determined a 1.05 Å crystal structure of an intermolecular pUG G4, which reveals an eight stranded G4 dimer involving 48 nucleotides, 7 different G and U quartet conformations, 7 coordinated potassium ions, 8 sodium ions and a buried water molecule. A comparison of the intermolecular pUG G4 and intramolecular pUG fold structures provides insights into the molecular basis for G4 handedness and illustrates how a simple dinucleotide repeat sequence can form complex structures with diverse topologies.

Multiple Regulatory Mechanisms Synergistically Control the Soluble Expression of CsCE for Enhanced Enzymatic Productivity of Lactulose in E. coli.

The limited expression of cellobiose 2-epimerase poses a significant constraint on the industrial enzymatic production of lactulose. Extensive modifications to the expression cassette offer a means to enhance the yield of recombinant proteins. In this study, an integrated strategy, combining individual and collaborative approaches, is proposed to fine-tune each stage of the CsCE overexpression program. This strategy involves the multidimensional integration of standardized genetic elements at various levels, including transcription, translation, folding, and three-dimensional structure. The volumetric activity of the final recombinant strain was markedly increased by 12-fold compared to the wild-type strain, reaching 2260.62 U/L. The protein expression in the newly developed high-yield recombinant strain exhibited a significant enhancement, with a higher proportion of soluble protein compared to that of inclusion bodies. Our findings offer insights into the multifaceted synergistic regulation of protein expression processes, holding promising implications for the production of heterologous recombinant proteins.

A comprehensive study of a new cylindrical flexible Miura-Ori origami: Kinematics, FEA, and fatigue assessments

In this paper, we introduced a new cylindrical flexible structure incorporating a multi-story origami based on the Miura-Ori pattern and mathematized kinematics. Our experiments validated the finite element simulations as well as the feasibility of the kinematics. We developed a fabrication technique and made a sample to study the feasibility and applicability of the designed structure. Subsequently, we set up a finite element simulation that mirrors the prepared sample. A series of axial folding experiments were carried out to validate the FEM simulation. We have also derived kinematic formulations relating the structure's height, folding angle, and radius. To model and analyze the folding and unfolding mechanisms of this innovative structure, we employed Finite Element Analysis (FEA). In our study, we investigate four main geometric parameters that significantly influence our design's characteristics: crease thickness, number of stories, crease width, and number of vertexes in one story. By varying these design parameters, we systematically examine their impact on key structural attributes such as fatigue life, folding force, and energy absorption. Our findings indicate that reduced crease thickness correlates with lower von-Mises stress and strain, significantly extending the origami design's lifespan. Through our analysis, we identify specific parameter values that maximize fatigue life, ensuring the structural integrity and longevity of the design. Our findings show that increasing the crease width extends the fatigue life dramatically. In conclusion, we summarize the influence of each design parameter on the structure's behavioral characteristics in a comprehensive approach.

Exploring phosphorene-protein interactions: An integrated computational and spectroscopic investigation

Among 2D materials, exfoliated black phosphorus (or phosphorene) shows great promise for applications in biological domains. However, despite its performances, little is known about the intricate and dynamic interactions that this material can form with proteins. This increases the risk of off-target effects and adds complexity in designing phosphorene-based devices with tailored properties. In this study, we present a straightforward and easily implementable pipeline that integrates spectroscopies with Molecular Dynamics simulations to explore the dynamic interplay between phosphorene and a protein system. Using lysozyme as a deeply investigated reference protein, we employed two theoretical protein models with unique secondary structure folds to increase the descriptive power of the approach and disentangle the complexity and variability of experimental data into a few primary drivers of protein-phosphorene interactions. Our results show that the 2D material does not significantly alter the protein structure, but the observed conformational changes are influenced by the secondary fold. Indeed, while the beta structure interacts mainly through unfolded regions, the alpha fold favours phosphorene binding through structured clusters of residues, leading to more significant structural and dynamic perturbations. By utilizing this pipeline, we have gained valuable insights into the molecular recognition mechanism of phosphorene, enhancing the development of improved phosphorene-based devices. In addition, our methodology offers potential for further applications in biomedicine to characterise interfaces between other 2D (nano)materials and biological entities.

Kinetic and Structural Studies of the Plastidial Solanum tuberosum Phosphorylase.

Kinetics and structural studies of the plastidial Solanum tuberosum phosphorylase (stPho1) revealed that the most active form of the enzyme (stPho1ΔL78) is composed by two segments generated by proteolytic degradation of an approximately 65-residue-long peptide (L78) approximately in the middle of the stPho1 primary structure. stPho1ΔL78 is 1.5 times more active than the nonproteolyzed enzyme in solution and shows stronger specificity for glycogen, α-d-glucose, caffeine, and β-cyclodextrin than stPho1. The crystal structure of stPho1ΔL78 has been resolved at 2.2 Å resolution and revealed similarities and differences with the mammalian enzymes. The structural fold is conserved as is the active site, while other binding sites such as the inhibitor, the glycogen storage, the quercetin, and the allosteric are not. The binding of α-d-glucose, caffeine, and β-cyclodextrin to stPho1 has been studied by X-ray crystallography and revealed significant differences from those of the mammalian phosphorylases. As stPho1 is capable of catalyzing both starch synthesis and degradation, our studies suggest that the direction of stPho1 activity is regulated by the proteolytic degradation of the L78 peptide.

Polymer Physics Models Reveal Structural Folding Features of Single-Molecule Gene Chromatin Conformations.

Here, we employ polymer physics models of chromatin to investigate the 3D folding of a 2 Mb wide genomic region encompassing the human LTN1 gene, a crucial DNA locus involved in key cellular functions. Through extensive Molecular Dynamics simulations, we reconstruct in silico the ensemble of single-molecule LTN1 3D structures, which we benchmark against recent in situ Hi-C 2.0 data. The model-derived single molecules are then used to predict structural folding features at the single-cell level, providing testable predictions for super-resolution microscopy experiments.

Structural style & kinematic analysis of deformation in the northern Dezful Embayment, Zagros Fold-Thrust Belt, SW Iran

A comprehensive understanding of structure and kinematic characteristics of fold and thrust belts provides significant information for hydrocarbon exploration and production. The kinematic evolution and structural style of three subsurface oilfields, Zeloi, Lali and Karun, in the northern part of the Dezful Embayment, a significant petroleum province of the Zagros Fold-Thrust Belt in SW Iran, were investigated using 2D seismic data. Interpretation of the 2D seismic profiles with up to 5 km depth consisting of upper Jurassic to Pleistocene sediments and the balanced cross-sections constructed revealed diverse geometries and kinematics along the oilfields. These results demonstrate that the structural style of the oilfields was controlled by the Gachsaran and Dashtak formations as upper and middle detachment levels, respectively. Tear faults affected the along-strike variations in structural style of the oilfields. Based on the analysis of growth strata, folding evolved as limbs rotated and hinges migrated, beginning in the mid-Miocene. During the later stages of deformation, the initial detachment folds transformed into fault-bend folds.

Genetic Variation in Protein Structure and Phylogenetic Relationships of West African Dwarf Sheep: An Investigative Analysis

This research aimed to ascertain the secondary and tertiary protein structure variations in the CD14 gene of the West African Dwarf sheep (Ovis aries) from Cross River and Akwa Ibom States, Nigeria. The research used forty samples (20 from each state) of West African Dwarf sheep (WAD) from farms and markets in the two States. One ml of blood was collected from each sheep through jugular venipuncture for DNA extraction. The region of CD14 of African Dwarf sheep was amplified by the Polymerase Chain Reaction (PCR) technique using specific sets of forward and reverse primers. The percentage of alpha helix in the secondary protein structure of WAD from AKS was higher than that detected in CRS. For the extended strand, the highest percentage was recorded from AKS17 (15.85%), while the lowest was from CRS14 (14.44%). Random coils had the highest percentage in the secondary protein structure and were generally similar among the selected WAD. The folding of the tertiary protein structure was generally similar among the WAD showing a close relationship in the activities of CD14 among the sheep. There was a close relationship among the WAD from Akwa Ibom and Cross River States based on CD14 gene variation.

Aerodynamic Analysis and Simulation for a Z-Shaped Folding Wing UAV

The potential of folding wing unmanned aerial vehicles (UAVs) in enhancing the adaptability of complex missions is substantial. Analyzing the aerodynamic characteristics of these UAVs is vital for optimizing the design of their folding wing configuration and airfoil shape. In this study, a model of a Z-shaped folding wing UAV was established with the objective of enhancing UAV maneuverability. An integrated vortex lattice method (VLM) was employed to analyze the aerodynamic characteristics of the Z-shaped folding wing and numerical simulations were utilized to validate the obtained results, which prove the effectiveness of the integrated VLM in studying aerodynamic characteristics at a high angle of attack of the Z-shaped folding wing UAV. This is a new way to reduce computation time while maintaining accuracy to calculate such complex folding structures in high fidelity. Furthermore, the Z-shaped folding wing configuration demonstrates enhanced maneuverability at high angles of attack, and a Simulink flight simulation model based on the aerodynamic coefficients of the Z-shaped folding wing verifies this property. This analysis can properly predict the post-stall characteristics for the Z-shaped folding wing UAV to ensure the safety of the vehicle during flight.

Fused radical SAM and αKG-HExxH domain proteins contain a distinct structural fold and catalyse cyclophane formation and β-hydroxylation.

Two of nature's recurring binding motifs in metalloproteins are the CxxxCxxC motif in radical SAM enzymes and the 2-His-1-carboxylate motif found both in zincins and α-ketoglutarate and non-haem iron enzymes. Here we show the confluence of these two domains in a single post-translational modifying enzyme containing an N-terminal radical S-adenosylmethionine domain fused to a C-terminal 2-His-1-carboxylate (HExxH) domain. The radical SAM domain catalyses three-residue cyclophane formation and is the signature modification of triceptides, a class of ribosomally synthesized and post-translationally modified peptides. The HExxH domain is a defining feature of zinc metalloproteases. Yet the HExxH motif-containing domain studied here catalyses β-hydroxylation and is an α-ketoglutarate non-haem iron enzyme. We determined the crystal structure for this HExxH protein at 2.8 Å, unveiling a distinct structural fold, thus expanding the family of α-ketoglutarate non-haem iron enzymes with a class that we propose to name αKG-HExxH. αKG-HExxH proteins represent a unique family of ribosomally synthesized and post-translationally modified peptide modifying enzymes that can furnish opportunities for genome mining, synthetic biology and enzymology.

Structural definition of pseudorabies virus dUTPase reveals a novel folding dimer in the herpesvirus family

The pseudorabies virus (PRV) causes severe and fatal acute respiratory disease in pigs. During PRV proliferation, the enzyme deoxyuridine 5′-triphosphate nucleotide hydrolase (dUTPase) plays a pivotal role in maintaining a low dUTP/dTTP ratio, thereby ensuring the accuracy of viral DNA replication. However, its structure and catalytic mechanisms have not been fully elucidated. Here, we report the crystal structure of PRV dUTPase at a 2.24 Å resolution and demonstrate an unprecedented dimeric architecture, with a conserved enzyme activity center of the herpesvirus family. The enzyme activity center is located in a cavity between the two domains, forming a pocket for binding substrate dUMP and magnesium ions. Remarkably, the exquisite interface of the dimer is primarily composed of four antiparallel β-sheets, which form 11 hydrogen bonds between the residues P33-V36 and R242-A248 to maintain protein stability. To the best of our knowledge, this is the first report demonstrating that dUTPase exists as a dimer in the herpesvirus family. These findings not only present a novel fold dimeric structure but also deepen the scope of our comprehension of structural diversity in dUTPase family.

Quantitative Characterization of RCA-Based DNA Hydrogels - Towards Rational Materials Design.

DNA hydrogels hold significant promise for biomedical applications and can be synthesized through enzymatic Rolling Circle Amplification (RCA). Due to the exploratory nature of this emerging field, standardized RCA protocols specifying the impact of reaction parameters are currently lacking. This study varied template sequences and reagent concentrations, evaluating RCA synthesis efficiency and hydrogel mechanical properties through quantitative PCR (qPCR) and indentation measurements, respectively. Primer concentration and stabilizing additives showed minimal impact on RCA efficiency, while changes in polymerase and nucleotide concentrations had a stronger effect. Concentration of the circular template exerted the greatest influence on RCA productivity. An exponential correlation between hydrogel viscosity and DNA amplicon concentration was observed, with nucleobase sequence significantly affecting both amplification efficiency and material properties, particularly through secondary structures. This study suggests that combining high-throughput experimental methods with structural folding prediction offers a viable approach for systematically establishing structure-property relationships, aiding the rational design of DNA hydrogel material systems.

Coronal energy release by MHD avalanches

Context. Magnetohydrodynamic (MHD) instabilities, such as the kink instability, can trigger the chaotic fragmentation of a twisted magnetic flux tube into small-scale current sheets that dissipate as aperiodic impulsive heating events. In turn, the instability could propagate as an avalanche to nearby flux tubes and lead to a nanoflare storm. Our previous work was devoted to related 3D MHD numerical modeling, which included a stratified atmosphere from the solar chromosphere to the corona, tapering magnetic field, and solar gravity for curved loops with the thermal structure modelled by plasma thermal conduction, along with optically thin radiation and anomalous resistivity for 50 Mm flux tubes. Aims. Using 3D MHD modeling, this work addresses predictions for the extreme-ultraviolet (EUV) imaging spectroscopy of such structure and evolution of a loop, with an average temperature of 2–2.5 MK in the solar corona. We set a particular focus on the forthcoming MUSE mission, as derived from the 3D MHD modeling. Methods. From the output of the numerical simulations, we synthesized the intensities, Doppler shifts, and non-thermal line broadening in 3 EUV spectral lines in the MUSE passbands: Fe IX 171 Å, Fe XV 284 Å, and Fe XIX 108 Å, emitted by ∼1 MK, ∼2 MK, and ∼10 MK plasma, respectively. These data were detectable by MUSE, according to the MUSE expected pixel size, temporal resolution, and temperature response functions. We provide maps showing different view angles (front and top) and realistic spectra. Finally, we discuss the relevant evolutionary processes from the perspective of possible observations. Reults. We find that the MUSE observations might be able to detect the fine structure determined by tube fragmentation. In particular, the Fe IX line is mostly emitted at the loop footpoints, where we might be able to track the motions that drive the magnetic stressing and detect the upward motion of evaporating plasma from the chromosphere. In Fe XV, we might see the bulk of the loop with increasing intensity, with alternating filamentary Doppler and non-thermal components in the front view, along with more defined spots in the topward view. The Fe XIX line is very faint within the chosen simulation parameters; thus, any transient brightening around the loop apex may possibly be emphasized by the folding of sheet-like structures, mainly at the boundary of unstable tubes. Conclusions. In conclusion, we show that coronal loop observations with MUSE can pinpoint some crucial features of MHD-modeled ignition processes, such as the related dynamics, helping to identify the heating processes.

Theta-curves in proteins.

We study and characterize the topology of connectivity circuits observed in natively folded protein structures whose coordinates are deposited in the Protein Data Bank (PDB). Polypeptide chains of some proteins naturally fold into unique knotted configurations. Another kind of nontrivial topology of polypeptide chains is observed when, in addition to covalent bonds connecting consecutive amino acids in polypeptide chains, one also considers disulfide and ionic bonds between non-consecutive amino acids. Bonds between non-consecutive amino acids introduce bifurcation points into connectivity circuits defined by bonds between consecutive and nonconsecutive amino acids in analyzed proteins. Circuits with bifurcation points can form θ-curves with various topologies. We catalog here the observed topologies of θ-curves passing through bridges between consecutive and non-consecutive amino acids in studied proteins.

Bridging the Gap between Sequence and Structure Classifications of Proteins with AlphaFold Models

Classification of protein domains based on homology and structural similarity serves as a fundamental tool to gain biological insights into protein function. Recent advancements in protein structure prediction, exemplified by AlphaFold, have revolutionized the availability of protein structural data. We focus on classifying about 9000 Pfam families into ECOD (Evolutionary Classification of Domains) by using predicted AlphaFold models and the DPAM (Domain Parser for AlphaFold Models) tool. Our results offer insights into their homologous relationships and domain boundaries. More than half of these Pfam families contain DPAM domains that can be confidently assigned to the ECOD hierarchy. Most assigned domains belong to highly populated folds such as Immunoglobulin-like (IgL), Armadillo (ARM), helix-turn-helix (HTH), and Src homology 3 (SH3). A large fraction of DPAM domains, however, cannot be confidently assigned to ECOD homologous groups. These unassigned domains exhibit statistically different characteristics, including shorter average length, fewer secondary structure elements, and more abundant transmembrane segments. They could potentially define novel families remotely related to domains with known structures or novel superfamilies and folds. Manual scrutiny of a subset of these domains revealed an abundance of internal duplications and recurring structural motifs. Exploring sequence and structural features such as disulfide bond patterns, metal-binding sites, and enzyme active sites helped uncover novel structural folds as well as remote evolutionary relationships. By bridging the gap between sequence-based Pfam and structure-based ECOD domain classifications, our study contributes to a more comprehensive understanding of the protein universe by providing structural and functional insights into previously uncharacterized proteins.

High-performance flexible pressure sensors with bionic dome-shaped fold structures inspired by crocodile skin

Flexible pressure sensors have drawn substantial community interest as a result of the advancement of contemporary technology. Inspired by the unique sensory organs of crocodiles, an ordered bionic dome-shaped fold structure array was prepared using a facile process in this paper, and electrical experiments confirmed the enhancement of the capabilities of piezoresistive flexible pressure sensors. The results revealed that this type of flexible pressure sensor presented a sensitivity of 8.87 kPa−1, could withstand a maximum pressure of 60 kPa, a minimum detection pressure of 10 Pa, a response time of 70 ms, and maintained stable performance under 5000 repeated presses. In addition, the flexible pressure sensors based on dome-shaped fold structure arrays have been successfully applied for human signal detection and plantar pressure monitoring system. The current research has significant implications for the manufacturing and practical applications of highly effective flexible pressure sensors.

Tissue-scale in vitro epithelial wrinkling and wrinkle-to-fold transition

Although epithelial folding is commonly studied using in vivo animal models, such models exhibit critical limitations in terms of real-time observation and independent control of experimental parameters. Here, we develop a tissue-scale in vitro epithelial bilayer folding model that incorporates an epithelium and extracellular matrix (ECM) hydrogel, thereby emulating various folding structures found in in vivo epithelial tissue. Beyond mere folding, our in vitro model realizes a hierarchical transition in the epithelial bilayer, shifting from periodic wrinkles to a single deep fold under compression. Experimental and theoretical investigations of the in vitro model imply that both the strain-stiffening of epithelium and the poroelasticity of ECM influence the folded structures of epithelial tissue. The proposed in vitro model will aid in investigating the underlying mechanism of tissue-scale in vivo epithelial folding relevant to developmental biology and tissue engineering.

Exploring natural product library as potential target against sterol C-24 methyltransferase protein of Leishmania donovani

Natural product offers an ocean of biologically active compounds that have diverse functionality. Thus, the present study aims for the exploration of natural product molecules for their leishmanicidal potency. Primary evaluation at 50 µM concentration revealed that out of 560 molecules, 38 compounds demonstrated a percentage killing of >50%. Next, the dose-dependent investigation showed that six active hits displayed the IC50 value ranging from 0.47 to 14.2 µM. Further, the molecular docking analysis using the alpha fold structure of Sterol C-24 methyltransferase of Leishmania donovani (LdSMT) (an enzyme absent in mammalian host) unveiled the strong binding affinity with top two hits namely shatavarin IV (−7.9 kcal/mol) and 6-methoxydihydrochelerythrine (−7.6 kcal/mol). Also, in silico studies were supported by the alterations in ergosterol content in the parasites treated with these two potent hits. In conclusion, our study suggests that the two potent hits inhibit the Leishmania parasite growth by hindering sterol biosynthesis.