641 research outputs found
Molecular jamming - the cystine slipknot mechanical clamp in all-atom simulations
A recent survey of 17 134 proteins has identified a new class of proteins
which are expected to yield stretching induced force-peaks in the range of 1
nN. Such high force peaks should be due to forcing of a slip-loop through a
cystine ring, i.e. by generating a cystine slipknot. The survey has been
performed in a simple coarse grained model. Here, we perform all-atom steered
molecular dynamics simulations on 15 cystine knot proteins and determine their
resistance to stretching. In agreement with previous studies within a coarse
grained structure based model, the level of resistance is found to be
substantially higher than in proteins in which the mechanical clamp operates
through shear. The large stretching forces arise through formation of the
cystine slipknot mechanical clamp and the resulting steric jamming. We
elucidate the workings of such a clamp in an atomic detail. We also study the
behavior of five top strength proteins with the shear-based mechanostability in
which no jamming is involved. We show that in the atomic model, the jamming
state is relieved by moving one amino acid at a time and there is a choice in
the selection of the amino acid that advances the first. In contrast, the
coarse grained model also allows for a simultaneous passage of two amino acids
Mechanical Stretching of Proteins: Calmodulin and Titin
Mechanical unfolding of several domains of calmodulin and titin is studied
using a Go-like model with a realistic contact map and Lennard-Jones contact
interactions. It is shown that this simple model captures the experimentally
observed difference between the two proteins: titin is a spring that is tough
and strong whereas calmodulin acts like a weak spring with featureless
force-displacement curves. The difference is related to the dominance of the
alpha secondary structures in the native structure of calmodulin. The tandem
arrangements of calmodulin unwind simultaneously in each domain whereas the
domains in titin unravel in a serial fashion. The sequences of contact events
during unraveling are correlated with the contact order, i.e. with the
separation between contact making amino acids along the backbone in the native
state. Temperature is found to affect stretching in a profound way.Comment: To be published in a special bio-issue of Physica A; 14 figure
Domain-domain interactions in Filamin A (16-23) impose a hierarchy of unfolding forces
The quaternary structure of Filamin A (FLNa) 16-23 was recently shown to
exhibit multiple domain-domain interactions that lead to a propeller-like
construction. Here we present single molecule force spectroscopy experiments to
show a wide variety of mechanical responses of this molecule and compare it
with its linear counterpart FLNa 1-8. The compact structure of FLNa 16-23 leads
to a broad distribution of rupture forces and end-to-end lengths in the
force-extension mode and multiple unraveling timescales in the force-clamp
mode. Moreover, a subset of force-extension trajectories reveals a mechanical
hierarchy in which the rupture of domain-domain interactions at high forces
(200 pN) liberates the unfolding of individual domains at low forces (100 pN).
This mechanism may also explain the order of magnitude difference in the rates
of the biexponential fits to the distribution of unfolding dwell times under
force-clamp. Overall, FLNa 16-23 under a force of 100 pN is more compliant than
the linear FLNa 1-8. Since a physiological role of FLNa is to crosslink actin
filaments, this range of responses allows it to accommodate a broad spectrum of
forces exerted by the cell and its environment
SingleâMolecule Mechanical Unfolding of Amyloidogenic β2âMicroglobulin: The ForceâSpectroscopy Approach
The recombinant production of a novel chimeric polyprotein is described. The new protein contains either wild-type β2-microglobulin (β2m) or its truncated variant (ÎN6 β2m) (see picture). Structural characterization is achieved by means of single-molecule force spectroscopy studies of specific β2m regions which could be involved in amyloidogenesis
BSDB: the biomolecule stretching database
We describe the Biomolecule Stretching Data Base that has been recently set up at http://www.ifpan.edu.pl/BSDB/. It provides information about mechanostability of proteins. Its core is based on simulations of stretching of 17â134 proteins within a structure-based model. The primary information is about the heights of the maximal force peaks, the forceâdisplacement patterns, and the sequencing of the contact-rupturing events. We also summarize the possible types of the mechanical clamps, i.e. the motifs which are responsible for a protein's resistance to stretching
Mechanical Strength of 17 134 Model Proteins and Cysteine Slipknots
A new theoretical survey of proteins' resistance to constant speed stretching
is performed for a set of 17 134 proteins as described by a structure-based
model. The proteins selected have no gaps in their structure determination and
consist of no more than 250 amino acids. Our previous studies have dealt with
7510 proteins of no more than 150 amino acids. The proteins are ranked
according to the strength of the resistance. Most of the predicted top-strength
proteins have not yet been studied experimentally. Architectures and folds
which are likely to yield large forces are identified. New types of potent
force clamps are discovered. They involve disulphide bridges and, in
particular, cysteine slipknots. An effective energy parameter of the model is
estimated by comparing the theoretical data on characteristic forces to the
corresponding experimental values combined with an extrapolation of the
theoretical data to the experimental pulling speeds. These studies provide
guidance for future experiments on single molecule manipulation and should lead
to selection of proteins for applications. A new class of proteins, involving
cystein slipknots, is identified as one that is expected to lead to the
strongest force clamps known. This class is characterized through molecular
dynamics simulations.Comment: 40 pages, 13 PostScript figure
Deconvolution of dynamic mechanical networks
Time-resolved single-molecule biophysical experiments yield data that contain
a wealth of dynamic information, in addition to the equilibrium distributions
derived from histograms of the time series. In typical force spectroscopic
setups the molecule is connected via linkers to a read-out device, forming a
mechanically coupled dynamic network. Deconvolution of equilibrium
distributions, filtering out the influence of the linkers, is a straightforward
and common practice. We have developed an analogous dynamic deconvolution
theory for the more challenging task of extracting kinetic properties of
individual components in networks of arbitrary complexity and topology. Our
method determines the intrinsic linear response functions of a given molecule
in the network, describing the power spectrum of conformational fluctuations.
The practicality of our approach is demonstrated for the particular case of a
protein linked via DNA handles to two optically trapped beads at constant
stretching force, which we mimic through Brownian dynamics simulations. Each
well in the protein free energy landscape (corresponding to folded, unfolded,
or possibly intermediate states) will have its own characteristic equilibrium
fluctuations. The associated linear response function is rich in physical
content, since it depends both on the shape of the well and its diffusivity---a
measure of the internal friction arising from such processes like the transient
breaking and reformation of bonds in the protein structure. Starting from the
autocorrelation functions of the equilibrium bead fluctuations measured in this
force clamp setup, we show how an experimentalist can accurately extract the
state-dependent protein diffusivity using a straightforward two-step procedure.Comment: 9 pages, 3 figures + supplementary material 14 pages, 4 figure
Mechanical unfolding of RNA: From hairpins to structures with internal multiloops
Mechanical unfolding of RNA structures, ranging from hairpins to ribozymes,
using laser optical tweezer (LOT) experiments have begun to reveal the features
of the energy landscape that cannot be easily explored using conventional
experiments. Upon application of constant force (), RNA hairpins undergo
cooperative transitions from folded to unfolded states whereas subdomains of
ribozymes unravel one at a time. Here, we use a self-organized polymer (SOP)
model and Brownian dynamics simulations to probe mechanical unfolding at
constant force and constant-loading rate of four RNA structures of varying
complexity. Our work shows (i) the response of RNA to force is largely
determined by the native structure; (ii) only by probing mechanical unfolding
over a wide range of forces can the underlying energy landscape be fully
explored.Comment: 26 pages, 6 figures, Biophys. J. (in press
Refolding upon force quench and pathways of mechanical and thermal unfolding of ubiquitin
The refolding from stretched initial conformations of ubiquitin (PDB ID:
1ubq) under the quenched force is studied using the Go model and the Langevin
dynamics. It is shown that the refolding decouples the collapse and folding
kinetics. The force quench refolding times scale as tau_F ~ exp(f_q*x_F/k_B*T),
where f_q is the quench force and x_F = 0.96 nm is the location of the average
transition state along the reaction coordinate given by the end-to-end
distance. This value is close to x_F = 0.8 nm obtained from the force-clamp
experiments. The mechanical and thermal unfolding pathways are studied and
compared with the experimental and all-atom simulation results in detail. The
sequencing of thermal unfolding was found to be markedly different from the
mechanical one. It is found that fixing the N-terminus of ubiquitin changes its
mechanical unfolding pathways much more drastically compared to the case when
the C-end is anchored. We obtained the distance between the native state and
the transition state x_UF=0.24 nm which is in reasonable agreement with the
experimental data.Comment: 35 pages, 15 figures, 1 tabl
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