scholarly journals Single-molecule mechanical unfolding kinetics of unmodified Saccharomyces cerevisiae tRNAPhe: a hint to the tRNA chaperone-tRNA interaction mechanism

2021 ◽  
Author(s):  
Wenzhao Liu ◽  
Luyi Feng ◽  
Wenpeng Zhu ◽  
Zhenyu Zhou ◽  
Ran Chen ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Optical Tweezers ◽  
Single Molecule ◽  
Interaction Mechanism ◽  
Dynamics Simulation ◽  
Stem Loop ◽  
Steered Molecular Dynamics Simulation ◽  
Dynamics Simulations ◽  
Unfolding Kinetics ◽  
Kinetics Of

The biological activity of tRNA is closely related to its mechanical folding properties. Although previous studies focused on the folding and unfolding mechanism of tRNA, its kinetics are largely unknown. In this study, combining optical tweezers and molecule dynamics simulations, we characterized the mechanical folding and unfolding processes of a single unmodified Saccharomyces cerevisiae tRNAphe. We identified the intermediates and pathways for tRNA mechanical folding and unfolding in the presence of Mg2+, discovering that the folding/unfolding kinetics of D stem-loop and T stem-loop but not the anti-codon stem-loop significantly affected by their upstream and downstream structures. The cooperative unfolding of the tRNA in the presence of Mg2+ lead to a large hysteresis between the folding and unfolding pathway, and such hysteresis and unfolding cooperativity are significantly reduced by lowering the Mg2+ concentration or mutating the nucleotides forming the 'elbow' structure. Moreover, both steered molecular dynamics simulation and optical tweezers experiment results support that, formation of tertiary interactions in the elbow region increases energy barriers of the mechanical unfolding pathway, including those in between intermediates, and determines the overall unfolding cooperativity. Our studies may shed light on the detailed tRNA chaperone mechanism of TruB and TrmA.

scholarly journals Kinetic and structural mechanism for DNA unwinding by a non-hexameric helicase

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sean P. Carney ◽  
Wen Ma ◽  
Kevin D. Whitley ◽  
Haifeng Jia ◽  
Timothy M. Lohman ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Single Molecule ◽  
Atp Hydrolysis ◽  
Variable Number ◽  
Structural Basis ◽  
Variable Step Size ◽  
Dna Unwinding ◽  
Step Size ◽  
Structural Mechanism ◽  
Dynamics Simulations

AbstractUvrD, a model for non-hexameric Superfamily 1 helicases, utilizes ATP hydrolysis to translocate stepwise along single-stranded DNA and unwind the duplex. Previous estimates of its step size have been indirect, and a consensus on its stepping mechanism is lacking. To dissect the mechanism underlying DNA unwinding, we use optical tweezers to measure directly the stepping behavior of UvrD as it processes a DNA hairpin and show that UvrD exhibits a variable step size averaging ~3 base pairs. Analyzing stepping kinetics across ATP reveals the type and number of catalytic events that occur with different step sizes. These single-molecule data reveal a mechanism in which UvrD moves one base pair at a time but sequesters the nascent single strands, releasing them non-uniformly after a variable number of catalytic cycles. Molecular dynamics simulations point to a structural basis for this behavior, identifying the protein-DNA interactions responsible for strand sequestration. Based on structural and sequence alignment data, we propose that this stepping mechanism may be conserved among other non-hexameric helicases.

Molecular dynamics simulation of displacement cascades in Cu and Ni: Thermal spike behavior

1989 ◽  
Vol 4 (3) ◽  
pp. 579-586 ◽  
Author(s):  
T. Diaz de la Rubia ◽  
R. S. Averback ◽  
Horngming Hsieh ◽  
R. Benedek
Keyword(s):  
Molecular Dynamics ◽  
Marked Effect ◽  
Dynamics Simulation ◽  
Primary State ◽  
Thermal Spike ◽  
Defect Production ◽  
Displacement Cascades ◽  
Dynamics Simulations ◽  
Atomic Mixing ◽  
Kinetics Of

Molecular dynamics simulations of energetic displacement cascades in Cu and Ni were performed with primary-knock-on-atom (PKA) energies up to 5 keV. The interatomic forces were represented by the Gibson II (Cu) and the Johnson-Erginsoy (Ni) potentials. Our results indicate that the primary state of damage produced by displacement cascades is controlled basically by two phenomena: replacement collision sequences during the ballistic phase, and melting and resolidification during the thermal spike. The thermal-spike phase is of longer duration and has a more marked effect in Cu than in Ni. Results for atomic mixing, defect production, and defect clustering are presented and compared with experiment. Simulations of “heat spikes” in these metals suggest a model for “cascade collapse” based on the regrowth kinetics of the molten cascade core.

scholarly journals Impact of an alpha helix and a cysteine–cysteine disulfide bond on the resistance of bacterial adhesion pili to stress

2021 ◽  
Vol 118 (21) ◽  
pp. e2023595118
Author(s):  
Joseph L. Baker ◽  
Tobias Dahlberg ◽  
Esther Bullitt ◽  
Magnus Andersson
Keyword(s):  
Optical Tweezers ◽  
Disulfide Bond ◽  
Alpha Helix ◽  
Covalent Bond ◽  
Structural Features ◽  
Dynamics Simulation ◽  
E Coli ◽  
Drag Forces ◽  
Steered Molecular Dynamics Simulation ◽  
Class 1

Escherichia coli express adhesion pili that mediate attachment to host cell surfaces and are exposed to body fluids in the urinary and gastrointestinal tracts. Pilin subunits are organized into helical polymers, with a tip adhesin for specific host binding. Pili can elastically unwind when exposed to fluid flow forces, reducing the adhesin load, thereby facilitating sustained attachment. Here we investigate biophysical and structural differences of pili commonly expressed on bacteria that inhabit the urinary and intestinal tracts. Optical tweezers measurements reveal that class 1a pili of uropathogenic E. coli (UPEC), as well as class 1b of enterotoxigenic E. coli (ETEC), undergo an additional conformational change beyond pilus unwinding, providing significantly more elasticity to their structure than ETEC class 5 pili. Examining structural and steered molecular dynamics simulation data, we find that this difference in class 1 pili subunit behavior originates from an α-helical motif that can unfold when exposed to force. A disulfide bond cross-linking β-strands in class 1 pili stabilizes subunits, allowing them to tolerate higher forces than class 5 pili that lack this covalent bond. We suggest that these extra contributions to pilus resiliency are relevant for the UPEC niche, since resident bacteria are exposed to stronger, more transient drag forces compared to those experienced by ETEC bacteria in the mucosa of the intestinal tract. Interestingly, class 1b ETEC pili include the same structural features seen in UPEC pili, while requiring lower unwinding forces that are more similar to those of class 5 ETEC pili.

scholarly journals Folding-Unfolding Kinetics of Skeletal-Muscle Titin Molecules Explored with Force-Clamp Optical Tweezers

2012 ◽  
Vol 102 (3) ◽  
pp. 577a
Author(s):  
Zsolt Martonfalvi ◽  
Pasquale Bianco ◽  
Miklos Kellermayer
Keyword(s):  
Skeletal Muscle ◽  
Optical Tweezers ◽  
Force Clamp ◽  
Unfolding Kinetics ◽  
Kinetics Of

scholarly journals Force Unfolding Kinetics of RNA Using Optical Tweezers. I. Effects of Experimental Variables on Measured Results

2007 ◽  
Vol 92 (9) ◽  
pp. 2996-3009 ◽  
Author(s):  
Jin-Der Wen ◽  
Maria Manosas ◽  
Pan T.X. Li ◽  
Steven B. Smith ◽  
Carlos Bustamante ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Unfolding Kinetics ◽  
Kinetics Of

scholarly journals Rad51 facilitates filament assembly of meiosis-specific Dmc1 recombinase

2020 ◽  
Vol 117 (21) ◽  
pp. 11257-11264 ◽  
Author(s):  
Wei-Hsuan Lan ◽  
Sheng-Yao Lin ◽  
Chih-Yuan Kao ◽  
Wen-Hsuan Chang ◽  
Hsin-Yi Yeh ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Single Molecule ◽  
Meiotic Recombination ◽  
Direct Evidence ◽  
Protein Protein Interaction ◽  
Filament Assembly ◽  
Tethered Particle Motion ◽  
Physical Interactions ◽  
Kinetics Of

Dmc1 recombinases are essential to homologous recombination in meiosis. Here, we studied the kinetics of the nucleoprotein filament assembly ofSaccharomyces cerevisiaeDmc1 using single-molecule tethered particle motion experiments and in vitro biochemical assay. ScDmc1 nucleoprotein filaments are less stable than the ScRad51 ones because of the kinetically much reduced nucleation step. The lower nucleation rate of ScDmc1 results from its lower single-stranded DNA (ssDNA) affinity, compared to that of ScRad51. Surprisingly, ScDmc1 nucleates mostly on the DNA structure containing the single-stranded and duplex DNA junction with the allowed extension in the 5′-to-3′ polarity, while ScRad51 nucleation depends strongly on ssDNA lengths. This nucleation preference is also conserved for mammalian RAD51 and DMC1. In addition, ScDmc1 nucleation can be stimulated by short ScRad51 patches, but not by EcRecA ones. Pull-down experiments also confirm the physical interactions of ScDmc1 with ScRad51 in solution, but not with EcRecA. Our results are consistent with a model that Dmc1 nucleation can be facilitated by a structural component (such as DNA junction and protein–protein interaction) and DNA polarity. They provide direct evidence of how Rad51 is required for meiotic recombination and highlight a regulation strategy in Dmc1 nucleoprotein filament assembly.

scholarly journals Force Unfolding Kinetics of RNA using Optical Tweezers. II. Modeling Experiments

2007 ◽  
Vol 92 (9) ◽  
pp. 3010-3021 ◽  
Author(s):  
M. Manosas ◽  
J.-D. Wen ◽  
P.T.X. Li ◽  
S.B. Smith ◽  
C. Bustamante ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Unfolding Kinetics ◽  
Kinetics Of

Molecular Dynamics Simulations of Dialkyl Carbocyanine Dyes in a DPPC Bilayer: Atomistic Insights Into Single Molecule Fluorescence

2007 ◽  
Author(s):  
Ramachandra Gullapalli ◽  
Melik Demirel ◽  
Peter J. Butler
Keyword(s):  
Molecular Dynamics ◽  
Single Molecule ◽  
Membrane Lipids ◽  
Dynamics Simulation ◽  
Spectroscopic Techniques ◽  
Carbocyanine Dyes ◽  
Lipid Dynamics ◽  
Membrane Perturbation ◽  
Dppc Bilayer ◽  
Dynamics Simulations

Our group uses di-alkyl carbocyanine dyes to detect force-induced membrane perturbation in mechanotransduction studies (Butler et al 2000). These dyes are also used extensively in single molecule spectroscopic techniques to infer dynamics of native membrane lipids. However, the precise distribution and orientation of the dye in a bilayer and how well the dye dynamics mimic native lipid dynamics are not known. Thus we present the results of a 40 nanosecond molecular dynamics simulation of a fully hydrated bilayer containing 0, 2, or 4 molecules of 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI-C18) and 128 molecules of dipalmitoylphosphatidyl choline (DPPC).

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