Complex Folding Research Articles

Thermal infrared remote-sensing detection of thermal information associated with faults: A case study in Western Sichuan Basin, China

Using enhanced land surface temperatures (LSTs) image retrieved from Landsat ETM+, this article shows that thermal information associated with faults have been detected. These anomalies may be provided by geothermal natural convection through faults and partially influence the ground surface thermal environment. The study area in southern segment of Longmen Mountains thrust belt of Western Sichuan Basin contains complex faults and folds with recent earthquake activity. In order to study the faults for future oil exploration, we use LST data retrieved from Landsat thermal infrared band to detect the thermal information associated with faults. The LSTs are enhanced by filtering out anthropogenic activity and influence land cover classes, and interpolating to contour map. The spatial patterns of the enhanced images revealed the spatial correspondence between the thermal information and the dip planes of faults when compared with the explanation profiles and geologic features obtained from the 3D-seismic geophysical data. The thermal-affected ranges calculated and the statistically significant of regression model also indicate the result that the thermal information located near the faults are consistent with the faults’ dip planes.

Subsalt Depth Seismic Imaging and Structural Interpretation in Dumre Area, Albania

The challenge of seismic exploration in fold and thrust belt settings is to optimize the depth seismic images of the deep structural objectives beneath a complex overburden that may show strong horizontal and vertical velocity variations. In such areas, the seismic image is frequently of poor quality and the depth models of deep layers is often false due to the perturbed propagation of seismic energy through the deforming lens of the overlying layers. A range of seismic processing tools, including post-stack and pre-stack depth migrations, are appropriate to predict the accurate geometry of deep target structures and to improve the building of a depth structural model.A strong combination of geological reasoning and depth seismic imaging processing can improve the understanding of the deep geological structures by reducing the uncertainties in depth geometrical and velocity model estimation. We propose an interpretative and iterative approach to the post stack depth migration method to guide the interpreter in the development of a reliable subsurface model. We have applied this approach during an exploration study in the Dumre area, located in the Ionian Basin (Albania) which is a complex fold and thrust belt. The main objectives of this study were to understand the failure of a former exploration well and to propose a new location for the potential closure of the carbonate structure. This subsalt imaging study aims at illustrating the improvements obtained by application of this integrated seismic imaging method especially in the evaluation of a subthrust prospect in a tectonically complex belt setting.

Kinetic Partitioning Mechanism Governs the Folding of the Third FnIII Domain of Tenascin-C: Evidence at the Single-Molecule Level

Statistical mechanics and molecular dynamics simulations proposed that the folding of proteins can follow multiple parallel pathways on a rugged energy landscape from unfolded state en route to their folded native states. Kinetic partitioning mechanism is one of the possible mechanisms underlying such complex folding dynamics. Here, we use single-molecule atomic force microscopy technique to directly probe the multiplicity of the folding pathways of the third fibronectin type III domain from the extracellular matrix protein tenascin-C (TNfn3). By stretching individual (TNfn3) 8 molecules, we forced TNfn3 domains to undergo mechanical unfolding and refolding cycles, allowing us to directly observe the folding pathways of TNfn3. We found that, after being mechanically unraveled and then relaxed to zero force, TNfn3 follows multiple parallel pathways to fold into their native states. The majority of TNfn3 fold into the native state in a simple two-state fashion, while a small percentage of TNfn3 were found to be trapped into kinetically stable folding intermediate states with well-defined three-dimensional structures. Furthermore, the folding of TNfn3 was also influenced by its neighboring TNfn3 domains. Complex misfolded states of TNfn3 were observed, possibly due to the formation of domain-swapped dimeric structures. Our studies revealed the ruggedness of the folding energy landscape of TNfn3 and provided direct experimental evidence that the folding dynamics of TNfn3 are governed by the kinetic partitioning mechanism. Our results demonstrated the unique capability of single-molecule AFM to probe the folding dynamics of proteins at the single-molecule level.

ARCHAEAN GOLD MINERALISATION DURING POST-OROGENIC EXTENSION IN THE NEW CONSORT GOLD MINE, BARBERTON GREENSTONE BELT, SOUTH AFRICA

The area around New Consort Gold Mine (NCGM) was complexly deformed during at least five distinct events named D1NC to D5NC. The D1NC to D3NC events involve progressive shearing and folding linked to the 3.25 to 3.23 Ga accretionary history of the Barberton greenstone belt (D1NC and D2NC), and to the subsequent ~3.1 Ga emplacement and doming of surrounding batholiths (D3NC). During D3NC the area experienced intense strain partitioning with the development of a network of shear zones that envelop structural domains characterised by locally unique deformation histories. Around NCGM, D3NC events involved early shearing (D3aNC) along the Consort Bar followed by two episodes of 600 m scale folding (D3bNC and D3cNC), resulting in a complex fold interference pattern. Pegmatites intruded during D3NC. The critical structures controlling gold mineralisation are D4NC extensional, 10 to 100 m scale, brittle-ductile shear fractures associated with kink bands and crenulation folds that formed concomitantly with gold mineralisation after all the D3NC structures had fully developed, marking a clearly separate event. This network of fractures is distributed along 100 to 200 m wide west-northwesterly, north-northwesterly and east-northeasterly trending corridors. The distribution of high-grade ore zones was controlled by the intersection orientation of D4NC fractures and suitable host lithologies, mainly the silicified hinge zones of D1NC folds and the siliceous Consort Bar. Because these host lithologies were complexly folded in D1NC-D3bNC-D3cNC fold interference patterns, the 3-D distribution of ore zones is highly discontinuous and complex. D5NC structures represent late reverse faults of unknown age, well developed in the nearby Albion Mine. Kinematic analyses and stress inversion using Bingham tensor solutions and a reduced moment tensorn method, of mineralised D4NC fractures along old gold workings at NCGM; consistently indicate a vertical σ1, and a horizontal northwesterly to southeasterly directed σ3, within an extensional stress regime. Results for the nearby Clutha, Albion and Woodstock mines are similar. This study suggests that gold mineralisation in the NCGM area can be linked to an extensional event that may have developed separately from the accretionary events shaping the craton, and may have coincided with deposition of the Dominion Group.

Unusual regioversatility of acetyltransferase Eis, a cause of drug resistance in XDR-TB

The emergence of multidrug-resistant and extensively drug-resistant (XDR) tuberculosis (TB) is a serious global threat. Aminoglycoside antibiotics are used as a last resort to treat XDR-TB. Resistance to the aminoglycoside kanamycin is a hallmark of XDR-TB. Here, we reveal the function and structure of the mycobacterial protein Eis responsible for resistance to kanamycin in a significant fraction of kanamycin-resistant Mycobacterium tuberculosis clinical isolates. We demonstrate that Eis has an unprecedented ability to acetylate multiple amines of many aminoglycosides. Structural and mutagenesis studies of Eis indicate that its acetylation mechanism is enabled by a complex tripartite fold that includes two general control non-derepressible 5 (GCN5)-related N-acetyltransferase regions. An intricate negatively charged substrate-binding pocket of Eis is a potential target of new antitubercular drugs expected to overcome aminoglycoside resistance.

Onshore record of the subduction of a crustal salient: example of the NW Borneo Wedge

Terra Nova, 23, 232–240, 2011AbstractSubduction of oceanic highs has been described in offshore accretionary wedges thanks to bathymetric and seismic data, and simulated in sand‐box models. Where documented, it is limited to oceanic subduction of small culminations (seamounts). However, tracing this phenomenon in ancient and/or uplifted belts remains a challenge. Based on structural, geomorphological and thermal analysis, we describe a subducted horst in the NW Borneo Wedge. This is evidenced by the occurrence of several geomorphological anomalies: a strong bend of the structural trend of the Rajang–Crocker Belt and an area with ′hummocky′ texture representing dismantled packages of sediments. Some specific structural features (large back and out‐of‐sequence thrusts in the internal zones, complex folds rooted on shear structures in the accretionary wedge) also occur in this central part of the NW Borneo Wedge. The central area of the related deformation zone records several thermal anomalies.Terra Nova, 00, 1–9, 2011

Flow of partially molten crust and the internal dynamics of a migmatite dome, Naxos, Greece

Migmatite domes are common in metamorphic core complexes. Dome migmatites deform in the partially molten or magmatic state and commonly record complex form surfaces, folds, and fabrics while units mantling the dome display a simpler geometry, typically formed by transposition during crustal extension. We use field observations and magnetic fabrics in the Naxos dome (Greece) to quantify the complex flow of anatectic crust beneath an extensional detachment system. The internal structure of the Naxos dome is characterized by second‐order domes (subdomes), pinched synforms, and curved lineation trajectories, which suggest that buoyancy‐driven flow participated in dome evolution. Subdomes broadly occur within two compartments that are separated by a steep, N‐S oriented, high‐strain zone. This pattern has been recognized in domes formed by polydiapirism and in models of isostasy‐dominated flow. The preferred model involves a combination of buoyancy‐ and isostasy‐driven processes: the Naxos dome may have been generated by regional N‐S extension that triggered convergent flow of partially molten crust at depth and the upwelling of anatectic migmatites within the dome. This pattern is complicated by gravitational instabilities and/or overturning of the high melt fraction crust leading to the growth of subdomes. As the migmatites within the Naxos dome reached a higher structural level, they were affected by regional top‐to‐the‐NNE kinematics of the detachment system. Dome formation therefore occurred by a combination of coeval and coupled processes: upper crustal extension, deep crust contraction during convergent flow of anatectic crust, diapirism and/or density‐driven crustal convection forming subdomes, and north directed detachment kinematics.

In Vitro Folding and Assembly of the Escherichia coli ATP-binding Cassette Transporter, BtuCD

Studies on membrane protein folding have focused on monomeric α-helical proteins and a major challenge is to extend this work to larger oligomeric membrane proteins. Here, we study the Escherichia coli (E. coli) ATP-binding cassette (ABC) transporter that imports vitamin B(12) (the BtuCD protein) and use it as a model system for investigating the folding and assembly of a tetrameric membrane protein complex. Our work takes advantage of the modular organization of BtuCD, which consists of two transmembrane protein subunits, BtuC, and two cytoplasmically located nucleotide-binding protein subunits, BtuD. We show that the BtuCD transporter can be re-assembled from both prefolded and partly unfolded, urea denatured BtuC and BtuD subunits. The in vitro re-assembly leads to a BtuCD complex with the correct, native, BtuC and BtuD subunit stoichiometry. The highest rates of ATP hydrolysis were achieved for BtuCD re-assembled from partly unfolded subunits. This supports the idea of cooperative folding and assembly of the constituent protein subunits of the BtuCD transporter. BtuCD folding also provides an opportunity to investigate how a protein that contains both membrane-bound and aqueous subunits coordinates the folding requirements of the hydrophobic and hydrophilic subunits.

Microsecond Subdomain Folding in Dihydrofolate Reductase

The characterization of microsecond dynamics in the folding of multisubdomain proteins has been a major challenge in understanding their often complex folding mechanisms. Using a continuous-flow mixing device coupled with fluorescence lifetime detection, we report the microsecond folding dynamics of dihydrofolate reductase (DHFR), a two-subdomain α/β/α sandwich protein known to begin folding in this time range. The global dimensions of early intermediates were monitored by Förster resonance energy transfer, and the dynamic properties of the local Trp environments were monitored by fluorescence lifetime detection. We found that substantial collapse occurs in both the locally connected adenosine binding subdomain and the discontinuous loop subdomain within 35 μs of initiation of folding from the urea unfolded state. During the fastest observable ∼ 550 μs phase, the discontinuous loop subdomain further contracts, concomitant with the burial of Trp residue(s), as both subdomains achieve a similar degree of compactness. Taken together with previous studies in the millisecond time range, a hierarchical assembly of DHFR—in which each subdomain independently folds, subsequently docks, and then anneals into the native conformation after an initial heterogeneous global collapse—emerges. The progressive acquisition of structure, beginning with a continuously connected subdomain and spreading to distal regions, shows that chain entropy is a significant organizing principle in the folding of multisubdomain proteins and single-domain proteins. Subdomain folding also provides a rationale for the complex kinetics often observed.

Identification of Multiple Folding Pathways of Monellin Using Pulsed Thiol Labeling and Mass Spectrometry

Protein folding reactions often display multiexponential kinetics of changes in intrinsic optical signals, as a manifestation of heterogeneity, either on one folding pathway or on multiple folding pathways. Delineating the origin of this heterogeneity is difficult because different coexisting structural forms of a protein cannot be easily distinguished by optical probes. In this study, the complex folding reaction of single-chain monellin has been investigated using a pulsed thiol labeling (SX) methodology in conjunction with mass spectrometry, which measures the kinetics of burial of a cysteine side chain thiol during folding. Because it can directly distinguish between unfolded and folded molecules and can measure the disappearance of the former during folding, the pulsed SX methodology is an ideal method for investigating whether multiple pathways are operative during folding. The kinetics of burial of the C42 thiol of monellin was observed to follow biexponential kinetics. To determine whether this was because the fast phase leads to the partial protection of the thiol group in all the molecules or to complete protection in only a fraction of the molecules, the duration and intensity of the labeling pulse were varied. The observation that the extent of labeling did not vary with the duration of the pulse cannot be explained by a simple sequential folding mechanism. Two parallel folding pathways are shown to be operative, with one leading to the formation of thiol-protective structure more rapidly than the other.

Physics-based de novo prediction of RNA 3D structures.

Current experiments on structural determination cannot keep up the pace with the steadily emerging RNA sequences and new functions. This underscores the request for an accurate model for RNA three-dimensional (3D) structural prediction. Although considerable progress has been made in mechanistic studies, accurate prediction for RNA tertiary folding from sequence remains an unsolved problem. The first and most important requirement for the prediction of RNA structure from physical principles is an accurate free energy model. A recently developed three-vector virtual bond-based RNA folding model ("Vfold") has allowed us to compute the chain entropy and predict folding free energies and structures for RNA secondary structures and simple pseudoknots. Here we develop a free energy-based method to predict larger more complex RNA tertiary folds. The approach is based on a multiscaling strategy: from the nucleotide sequence, we predict the two-dimensional (2D) structures (defined by the base pairs and tertiary contacts); based on the 2D structure, we construct a 3D scaffold; with the 3D scaffold as the initial state, we combine AMBER energy minimization and PDB-based fragment search to predict the all-atom structure. A key advantage of the approach is the statistical mechanical calculation for the conformational entropy of RNA structures, including those with cross-linked loops. Benchmark tests show that the model leads to significant improvements in RNA 3D structure prediction.

Protein disulfide isomerase isomerizes non‐native disulfide bonds in human proinsulin independent of its peptide‐binding activity

Protein disulfide isomerase (PDI) supports proinsulin folding as chaperone and isomerase. Here, we focus on how the two PDI functions influence individual steps in the complex folding process of proinsulin. We generated a PDI mutant (PDI-aba'c) where the b' domain was partially deleted, thus abolishing peptide binding but maintaining a PDI-like redox potential. PDI-aba'c catalyzes the folding of human proinsulin by increasing the rate of formation and the final yield of native proinsulin. Importantly, PDI-aba'c isomerizes non-native disulfide bonds in completely oxidized folding intermediates, thereby accelerating the formation of native disulfide bonds. We conclude that peptide binding to PDI is not essential for disulfide isomerization in fully oxidized proinsulin folding intermediates.

Folding Kinetics of the Cooperatively Folded Subdomain of the IκBα Ankyrin Repeat Domain

The ankyrin repeat (AR) domain of IκBα consists of a cooperative folding unit of roughly four ARs (AR1–AR4) and of two weakly folded repeats (AR5 and AR6). The kinetic folding mechanism of the cooperative subdomain, IκBα67-206, was analyzed using rapid mixing techniques. Despite its apparent architectural simplicity, IκBα67-206 displays complex folding kinetics, with two sequential on-pathway high-energy intermediates. The effect of mutations to or away from the consensus sequences of ARs on folding behavior was analyzed, particularly the GXTPLHLA motif, which have not been examined in detail previously. Mutations toward the consensus generally resulted in an increase in folding stability, whereas mutations away from the consensus resulted in decreased overall stability. We determined the free energy change upon mutation for three sequential transition state ensembles along the folding route for 16 mutants. We show that folding initiates with the formation of the interface of the outer helices of AR3 and AR4, and then proceeds to consolidate structure in these repeats. Subsequently, AR1 and AR2 fold in a concerted way in a single kinetic step. We show that this mechanism is robust to the presence of AR5 and AR6 as they do not strongly affect the folding kinetics. Overall, the protein appears to fold on a rather smooth energy landscape, where the folding mechanism conforms a one-dimensional approximation. However, we note that the AR does not necessarily act as a single folding element.

The Yeast Prion Case: Could There be a Uniform Concept Underlying Complex Protein Folding?

The Yeast Prion Case: Could There be a Uniform Concept Underlying Complex Protein Folding?

Bacterial proteins fold faster than eukaryotic proteins with simple folding kinetics

Protein domain frequency and distribution among kingdoms was statistically analyzed using the SCOP structural database. It appeared that among chosen protein domains with the best resolution, eukaryotic proteins more often belong to α-helical and β-structural proteins, while proteins of bacterial origin belong to α/β structural class. Statistical analysis of folding rates of 73 proteins with known experimental data revealed that bacterial proteins with simple kinetics (23 proteins) exhibit a higher folding rate compared to eukaryotic proteins with simple folding kinetics (27 proteins). Analysis of protein domain amino acid composition showed that the frequency of amino acid residues in proteins of eukaryotic and bacterial origin is different for proteins with simple and complex folding kinetics.

Unzip Single Protein Zippers using Optical Tweezers

Basic leucine zippers (bZIPs) are dimeric alpha-helical domains found in many transcription factors. They comprise coiled coil regions (zippers) that mediate specific protein dimerization, and basic regions that undergo disorder-to-order transitions to grip their cognate DNA binding sites. It remains unclear how folding of the two regions and binding of DNA are coupled. We investigated the detailed folding kinetics of a model bZIP domain derived from GCN4 transcriptional factor, using high-resolution optical tweezers. When pulled from the same amino- or carboxyl-terminals, the coiled coil alone folds and unfolds rapidly in a two-state manner, with little energy barrier. The folding kinetics of the full GCN4 bZIP domain is similar to the coiled coil alone. However, the presence of its binding site (AP1) induces a third new state corresponding to the bZIP-DNA complex. The probability of this new state critically depends upon the AP1 concentration, which reaches a maximum at an intermediate AP1 concentration. Results from our single-molecule manipulation experiments reveal a complex folding energy landscape even for a small protein domain that can be further tuned by their binding ligands.

Folding Studies of Beta-Strand-Containing Repeat Proteins Through Naturally-Occurring and Consensus-Designed Sequences

Repeat proteins, devoid of sequence-distant contacts observed in globular proteins, are ideal candidates for the dissection of local stability, nearest-neighbor contact parameters, cooperativity, and determination of folding energy landscapes. Recent studies on HEAT, TPR, Ankyrin, and Armadillo have contributed to the understanding of the folding of helical repeat proteins. Moreover, constructs composed of simplified “consensus” sequence representations of helical repeat proteins have been found to be well-behaved and highly stable, and facilitate energy dissection using simple nearest-neighbor models. In contrast, little progress has been made towards understanding the folding of β-sheet containing repeat. To determine the folding stability and cooperativity of these proteins, and to better understand the sequence determinants of structure and stability within these ubiquitous families, we have initiated studies on a series of LRR proteins of both naturally occurring and consensus-designed sequences. We find the LRR proteins PP32 and LC1 to be well-behaved, and to fold in a highly cooperative transition that is consistent with a two-state mechanism. NMR H/2H exchange shows the repeating β-strands on the concave surface of both proteins to be more protected than the rest of the molecules, and can be regarded as an exchange-resistant core, whereas the terminal caps and convex structural elements are more labile. However, truncations and sequence substitution demonstrate that the caps significantly influence stability and kinetics. To further simplify our analysis of β-sheet containing repeat protein folding, we designed consensus LRR sequences. On their own, these constructs are unfolded and/or aggregated. By fusing these consensus sequences with naturally occurring LRR protein YopM, we have obtained solublized, folded arrays that exhibit increased stability and drastically decreased unfolding and refolding rates with repeat number. Further studies are needed to dissect the complex folding pathways taken by these constructs.

Augmenting β-Augmentation: Structural Basis of How BamB Binds BamA and May Support Folding of Outer Membrane Proteins

β-Barrel proteins are frequently found in the outer membrane of mitochondria, chloroplasts and Gram-negative bacteria. In Escherichia coli, these proteins are inserted in the outer membrane by the Bam (β-barrel assembly machinery) complex, a multiprotein machinery formed by the β-barrel protein BamA and the four peripheral membrane proteins BamB, BamC, BamD and BamE. The periplasmic part of BamA binds prefolded β-barrel proteins by a β-augmentation mechanism, thereby stabilizing the precursors prior to their membrane insertion. However, the role of the associated proteins within the Bam complex remains unknown. Here, we describe the crystal structure of BamB, a nonessential component of the Bam complex. The structure shows a typical eight-bladed β-propeller fold. Two sequence stretches of BamB were previously identified to be important for interaction with BamA. In our structure, both motifs are located in close proximity to each other and contribute to a conserved region forming a narrow groove on the top of the propeller. Moreover, crystal contacts reveal two interaction modes of how BamB might bind unfolded β-barrel proteins. In the crystal lattice, BamB binds to exposed β-strands by β-augmentation, whereas peptide stretches rich in aromatic residues can be accommodated in hydrophobic pockets located at the bottom of the propeller. Thus, BamB could simultaneously bind to BamA and prefolded β-barrel proteins, thereby enhancing the folding and membrane insertion capability of the Bam complex.

New Scenarios of Protein Folding Can Occur on the Ribosome

Identifying and understanding the differences between protein folding in bulk solution and in the cell is a crucial challenge facing biology. Using Langevin dynamics, we have simulated intact ribosomes containing five different nascent chains arrested at different stages of their synthesis such that each nascent chain can fold and unfold at or near the exit tunnel vestibule. We find that the native state is destabilized close to the ribosome surface due to an increase in unfolded state entropy and a decrease in native state entropy; the former arises because the unfolded ensemble tends to behave as an expanded random coil near the ribosome and a semicompact globule in bulk solution. In addition, the unfolded ensemble of the nascent chain adopts a highly anisotropic shape near the ribosome surface and the cooperativity of the folding-unfolding transition is decreased due to the appearance of partially folded structures that are not populated in bulk solution. The results show, in light of these effects, that with increasing nascent chain length folding rates increase in a linear manner and unfolding rates decrease, with larger and topologically more complex folds being the most highly perturbed by the ribosome. Analysis of folding trajectories, initiated by temperature quench, reveals the transition state ensemble is driven toward compaction and greater native-like structure by interactions with the ribosome surface and exit vestibule. Furthermore, the diversity of folding pathways decreases and the probability increases of initiating folding via the N-terminus on the ribosome. We show that all of these findings are equally applicable to the situation in which protein folding occurs during continuous (non-arrested) translation provided that the time scales of folding and unfolding are much faster than the time scale of monomer addition to the growing nascent chain, which results in a quasi-equilibrium process. These substantial ribosome-induced perturbations to almost all aspects of protein folding indicate that folding scenarios that are distinct from those of bulk solution can occur on the ribosome.

Kinematic waves in polar firn stratigraphy

AbstractRadar studies of firn on the ice sheets have revealed complex folds on its internal layering that form from the interplay of snow accumulation and ice flow. A mathematical theory for these fold structures is presented, for the case where the radar cross section lies along the ice-flow direction and where the accumulation rate and ice-flow velocity are time-invariant. Our model, which accounts for firn densification, shows how ‘information’ (the depth and slope of isochrones) propagates on the radargram to govern its layer undulations. This leads us to discover universal rules behind the pattern of layer slopes on a distance–age domain and understand why the loci of layer-fold hinges curve, emerge and combine on the radargram to form closed loops that delineate areas of rising and plunging isochrones. We also develop a way of retrieving the accumulation rate distribution and layer ages from steady isochrone patterns. Analysis of a radargram from the onset zone of Bindschadler Ice Stream, West Antarctica, indicates that ice flow and accumulation rates have been steady there for the past ∼400 years, and that spatial anomalies in the latter are coupled to surface topography induced by ice flow over the undulating ice-stream bed. The theory provides new concepts for the morphological interpretation of radargrams.