Journal: bioRxiv

Article Title: Conformational dynamics of actin filaments crosslinked with alpha-actinin and their roles in suppressing cofilin-induced helical shortening and cluster formation

doi: 10.1101/2025.11.16.688667

Figure Lengend Snippet: ( A ) A longitudinal section profile illustrating for the analysis of two consecutive HHPs, the number of protomer pairs per HHP, and MADs in various actin filaments captured at different time points. The analytic methods used were developed in our previous study . ( B ) Still AFM images with time labels were shown for conditions with ATP present and after ATP depletion (primarily converted into ADP and inorganic phosphate in solution through ATPase activities of actin and S1) to observe the binding of S1 on actin filaments. As ATP levels gradually declined or nearly depleted due to ATP hydrolysis (ATP → ADP + Pi) during the actin-myosin reaction, S1 binding to actin filaments persisted for longer durations, enabling capture by HS-AFM at an imaging rate of 330 ms per frame. All actin filaments in the imaging field were aligned with the pointed end (PE) on the left and the barbed end (BE) on the right, reflecting the actual unipolar crosslinking configuration. As similarly done in our previous study by referring to the EM study , the polarity of actin filaments was confirmed by observing the tilted angles of S1, with its head pointing toward the PE, resembling an arrowhead along the filament. This characteristic orientation allowed differentiation between the PE and the BE. Representative yellow and cyan arrowheads indicate alpha-actinin and S1 molecules, respectively. Scale bars: 25 nm. Color scale: 0-12 nm. ( C ) Time-dependent changes in two consecutive HHPs, the number of protomer pairs per HHP, and MADs in actin filaments crosslinked by 50 nM alpha-actinin in the presence of 1 mM ATP, 0.1 mM CaCl₂, and 1 µM S1, but without MgCl 2 . Related to , Movie S1 .

Article Snippet: Movie S8 Molecular dynamics of unipolar-flexible crosslinking model obtained from all-atom MD simulations.

Techniques: Binding Assay, Imaging

Journal: bioRxiv

Article Title: Conformational dynamics of actin filaments crosslinked with alpha-actinin and their roles in suppressing cofilin-induced helical shortening and cluster formation

doi: 10.1101/2025.11.16.688667

Figure Lengend Snippet: ( A ) Experimental setup for HS-AFM observations of dynamic conformational changes in alpha-actinin-crosslinked actin filaments in the presence and absence of S1. ( B ) Representative AFM images of alpha-actinin-crosslinked actin filaments with and without S1. Yellow arrowheads representatively indicate alpha-actinin molecules; black arrowheads mark the longitudinal section points used to measure the HHP. Two parallelograms (ABCD and EFHG) represent the geometric framework used to measure the crosslinking angles of alpha-actinin and the crosslinking lengths between alpha-actinin-crosslinked actin filaments. ( C ) Histogram of HHP values for alpha-actinin-crosslinked actin filaments without S1. The mean HHP is 36.5 ± 2.8 nm (mean ± SD, N = 4270). F-buffer consists of 0.5 mM MgCl₂, 0.5 mM ATP, and 50 nM alpha-actinin at pH 6.8. ( D, E ) Ridgelines showing the distributions of alpha-actinin crosslinking angles and crosslinking lengths between filaments crosslinked with alpha-actinin, respectively. Noted that we focus solely on the four symmetric small crosslinking angles generated by two alpha-actinin molecules bridging two actin filaments, irrespective of their polarity and demonstrated in a parallelogram ABCD. Statistical significance was determined using a two-sample t-test , with differences considered significant at p ≤ 0.05. See Methods and for details.

Article Snippet: Movie S8 Molecular dynamics of unipolar-flexible crosslinking model obtained from all-atom MD simulations.

Techniques: Generated

Journal: bioRxiv

Article Title: Conformational dynamics of actin filaments crosslinked with alpha-actinin and their roles in suppressing cofilin-induced helical shortening and cluster formation

doi: 10.1101/2025.11.16.688667

Figure Lengend Snippet: ( A ) Experimental setup for HS-AFM observations cofilin-induced helical shortening, cofilin binding and clustering in bare actin filaments versus alpha-actinin-crosslinked-actin filaments. ( B ) Time-dependent changes in the peak heights and HHPs of actin filaments incubated with only 600 nM hs cofilin 1 and 1mM ATP. A ∼25% decrease in HHP and the formation of mature cofilin clusters on actin filaments in the absence of alpha-actinin served as positive controls, providing a basis for comparison with the cofilin clusters and HHP shown in C . Red and green arrowheads indicate the peaks in mature cofilin clusters and bare actin segments (served as negative controls), respectively. Time label indicates the time after adding cofilin. Scale bars: 25 nm. Color scale: 0-12 nm. ( C ) Time-dependent changes in the peak heights and HHPs of actin filaments incubated with 400 nM alpha-actinin and 1 mM ATP, and subsequently added with 600 nM cofilin. The cofilin binding and formation of mature cofilin clusters, and the shortening of HHP were significantly suppressed by binding and crosslinking of alpha-actinin. Red arrowheads denote the peaks in mature cofilin clusters within the supertwisted half helices of actin segments (cofilactin) without alpha-actinin, and blue arrowheads indicate those in immature cofilin clusters within the long half helices of actin segments crosslinked with alpha-actinin (cofilactin-actinin), respectively. A representative yellow arrowhead denotes alpha-actinin. Scale bars: 25 nm. Color scale: 0-13 nm. ( D ) Histograms of the peak height and HHP measured from various actin filaments with and without alpha-actinin binding and crosslinking, showing the presence of both mature and immature cofilin clusters. These clusters corresponded to short and normal HHPs, respectively. Notably, their categorization as mature or immature clusters was based on their differences in peak heights and length of corresponding HHPs, and their comparisons with bare actin (i.e., peak height of 8.6 ± 0.8 nm, HHP of 36.8 ± 4.3 nm), as reported in our previous studies . The mean ± SD heights and lengths are 10.7 ± 1.0 nm (N = 425) and 27.9 ± 3.0 nm (N= 300) for mature clusters of cofilactin segment without alpha-actinin, and 9.9 ± 1.1 nm (N = 2514) and 33.0 ± 4.6 nm (N = 2003) for immature clusters of cofilactin segment with alpha-actinin crosslinking (cofilactin-actinin). A two-population t-test ( p < 0.05) indicates that the differences in height and HHP distribution between the clusters of cofilactin segments without and with alpha-actinin crosslinking are significant ( p = 2.9×10 -38 for height, p = 8.5×10 -68 for HHP).

Article Snippet: Movie S8 Molecular dynamics of unipolar-flexible crosslinking model obtained from all-atom MD simulations.

Techniques: Binding Assay, Incubation, Comparison

Journal: bioRxiv

Article Title: Conformational dynamics of actin filaments crosslinked with alpha-actinin and their roles in suppressing cofilin-induced helical shortening and cluster formation

doi: 10.1101/2025.11.16.688667

Figure Lengend Snippet: ( A ) The unipolar and bipolar crosslinking models of the actin filaments with alpha-actinin. The unipolar crosslinking configuration occurs when the actin filaments share the same polarity, whereas the bipolar crosslinking configuration appears when the actin filaments align with opposite polarities. The quantitative analyses were performed using conformations obtained from all-atom MD simulations for 300 ns. ( B ) Quantitative analysis of the crosslinking lengths and alpha-actin crosslinking angles relative to the filament axis in the unipolar-flexible, bipolar-flexible, and bipolar-fixed models obtained from the MD simulations. ( C ) Analysis of the HHP, rise, and AD in the bare actin filaments and in the unipolar-flexible, bipolar-flexible, and bipolar-fixed models using the conformations derived from the MD simulations. ( D, E ) Characterization of the residue-level interactions between the alpha-actinin ABDs and actin protomers during crosslinking in the unipolar-flexible and bipolar-flexible models. In our model, if an atom of one residue (Cα) was in contact within 2 Å of an atom of another residue (Cα), these two residues were considered to be interacting. Our MD simulations identified key alpha-actinin–actin residue pairs with contact using probability density function (nm -1 ), which can be greater than 1 since there are two alpha-actinin ends interacting actin filaments. Thus, the maximum probability value is 2.

Article Snippet: Movie S8 Molecular dynamics of unipolar-flexible crosslinking model obtained from all-atom MD simulations.

Techniques: Derivative Assay, Residue

Journal: bioRxiv

Article Title: Conformational dynamics of actin filaments crosslinked with alpha-actinin and their roles in suppressing cofilin-induced helical shortening and cluster formation

doi: 10.1101/2025.11.16.688667

Figure Lengend Snippet: ( A, B ) Twisted and flattened conformational transitions between G-actin, F-actin, and C-actin structures have been previously linked to actin polymerization and depolymerization and cofilin clustering. Specifically, twisted G-actin polymerizes to form flattened actin filament protomers (F-actin form) and vice versa , while cofilin-induced clustering leads to twisted protomers (C-actin), and subsequently shortening the HHP in actin filament accompanied by a reduction in the number of protomers per HHP , , . Our current study deciphers that alpha-actinin binding and crosslinking help preserve the normal canonical helical structure of actin filaments, with actin protomers most likely adopting a flattened conformation (F-actin). This conformation facilitates S1 binding but prevents cofilin binding or clustering. In contrast, the twisted conformation of actin protomers induced by cofilin binding and clustering induces the shortening of helical twists, which supports cofilin’s cooperative binding and clustering , . We suggest that the process of cooperativity of cofilin binding and clustering likely follows the sequence: initial cofilin binding and clustering onto shortened ADP-bound HHP → protomer twisting → HHP shortening accompanied by a reduction in the number of protomers per HHP → cooperative binding and clustering. However, there is currently no atomic structural evidence to confirm whether the twisted actin protomers within the shortened bare helices could inhibit S1 binding, as suggested from previous fluorescence-based observation , despite the known mechanism of direct competition for binding sites between cofilin and S1 on actin filaments . This remains an area for further investigation in future studies. PE: pointed end; BE: barbed end. ( C ) Illustration depicting alpha-actinin-crosslinked actin filaments under the unipolar and bipolar conformations with normal and rigid HHPs resistant to cofilin binding and/or clustering, highlighting their distinct localizations and functions within cells. This Figure was created using BioRender.com. Related to .

Article Snippet: Movie S8 Molecular dynamics of unipolar-flexible crosslinking model obtained from all-atom MD simulations.

Techniques: Binding Assay, Sequencing, Fluorescence

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 2. Schematic diagram of the initial construction of the slit displacement model (with red representing CO2, green representing C4, blue representing C12, and purple representing C20).

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 7. Schematic of the quartz fracture model at a displacement velocity of 1 m/s (red: CO2, green: C4, blue: C12, and purple: C20): (a) 1000 ps; (b) 3000 ps; (c) 7000 ps; and (d) 8000 ps.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 8. Fingering phenomenon of CO2 during the displacement process in the quartz model (green: C4, blue: C12, and purple: C20): (a) 1000 ps; and (b) 4000 ps.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 10 presents the interaction energy between alkanes and the rock wall surface, as well as the interaction energy between CO2 and alkanes. The former reflects the adsorption strength between shale oil and the rock surface, where a greater interaction force indicates a more stable adsorption state. In cases where the interaction energy between shale oil and the rock wall is high, CO2 has difficulty displacing and separating the alkanes adsorbed on the pore walls, thereby destabilizing the displacement interface. The latter primarily represents the miscibility between CO2 and the shale oil components. A greater interaction energy between alkanes and CO2 facilitates miscibility, enabling a miscible displacement process that stabilizes the displacement front. The adsorption strength of alkane molecules on the pore wall is relatively weak. During the displacement process, the interaction between CO2 and alkanes plays a dominant role. After CO2 displaces the alkanes from the slit, the reduced interaction between alkanes and CO2 leads to the increase observed at the end of the curve in the simulation.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques: Adsorption

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 11. Schematic of final displacement results at different displacement velocities in the quartz model: (a) 1 m/s, (b) 2 m/s, (c) 3 m/s, and (d) 4 m/s (red: CO2, green: C4, blue: C12, and purple: C20).

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 12. X-direction density distribution of alkane molecules at different displacement velocities in the quartz model ((a): t = 100 ps, and (b): t = final time).

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 15 presents snapshot diagrams of the calcite slit model under a displacement velocity of 1 m/s. These snapshots allow for a visual analysis of the displacement charac- teristics of each alkane molecule. CO2 molecules continuously advance, displacing and peeling off alkane molecules adsorbed on the pore wall surface. In the early stages of displacement, depressions form between the alkane molecules, and an oil film adsorption phenomenon can be observed on the left side of the slit model. High molecular weight alkanes (C12 and C20) adsorbed on the pore wall surface hinder the advancement of CO2 along the wall and negatively affect the displacement of alkanes within the slit. CO2 continues to advance towards the center of the pore, and as it progresses, the number of alkane molecules adsorbed on the alkane wall increases. From Figure 16, it is evident that the alkane molecules on the left side are compressed by CO2, and the displacement front is progressively overcome. As a result, a clear forward motion (finger-like displacement) of CO2 is observed, indicating that the CO2 at the center of the pore is in an ineffective displacement state. Additionally, the finger-like displacement phenomenon observed in the calcite model is more pronounced than that in the quartz model, suggesting that more CO2 molecules in the calcite model are in an ineffective displacement state.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques: Adsorption, High Molecular Weight

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 15. Snapshot of the calcite slit model at a displacement velocity of 1 m/s (red: CO2, green: C4, blue: C12, and purple: C20): (a) 1000 ps; (b) 3000 ps; (c) 7000 ps; and (d) 8000 ps.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 16. Finger-like displacement phenomenon of CO2 during the displacement process in the calcite model (green: C4, blue: C12, and purple: C20). (a) 1000 ps; and (b) 4000 ps.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 18. Schematic of final displacement results at different displacement velocities in the calcite model: (a) 1 m/s, (b) 2 m/s, (c) 3 m/s, (d) 4 m/s (red: CO2, green: C4, blue: C12, and purple: C20).

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 19. X-direction density distribution of alkane molecules at different displacement velocities in the calcite model. ((a): t = 100 ps, (b): t = final time).

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques:

Journal: Energies

Article Title: Molecular Dynamics Simulation on the Mechanism of Shale Oil Displacement by Carbon Dioxide in Inorganic Nanopores

doi: 10.3390/en18020262

Figure Lengend Snippet: Figure 19 illustrates the density distribution of alkane molecules in the X-direction at four different displacement velocities. It can be clearly observed that as the displacement velocity decreases, the peak position of the alkane molecule density distribution in the X-direction shifts further, indicating a more widespread and concentrated distribution of alkanes at lower velocities. The distribution of C12 and C20 at all velocities shows a broader spread. In general, lower displacement velocities are more favorable for the CO2 displacement process. However, as the displacement velocity increases, the ability of CO2 to displace alkanes weakens, leading to a shorter displacement distance of alkanes in the X-direction, which is consistent with the findings in the quartz model.

Article Snippet: Dynamic Displacement Process Molecular dynamics simulations were conducted using the quartz fracture displacement model constructed earlier.

Techniques: