248 research outputs found
Jamming proteins with slipknots and their free energy landscape
Theoretical studies of stretching proteins with slipknots reveal a surprising
growth of their unfolding times when the stretching force crosses an
intermediate threshold. This behavior arises as a consequence of the existence
of alternative unfolding routes that are dominant at different force ranges.
Responsible for longer unfolding times at higher forces is the existence of an
intermediate, metastable configuration where the slipknot is jammed.
Simulations are performed with a coarsed grained model with further
quantification using a refined description of the geometry of the slipknots.
The simulation data is used to determine the free energy landscape (FEL) of the
protein, which supports recent analytical predictions.Comment: 5 page
Constructing a folding model for protein S6 guided by native fluctuations deduced from NMR structures
The diversity in a set of protein nuclear magnetic resonance (NMR) structures provides an estimate of native state fluctuations that can be used to refine and enrich structure-based protein models (SBMs). Dynamics are an essential part of a proteinâs functional native state. The dynamics in the native state are controlled by the same funneled energy landscape that guides the entire folding process. SBMs apply the principle of minimal frustration, drawn from energy landscape theory, to construct a funneled folding landscape for a given protein using only information from the native structure. On an energy landscape smoothed by evolution towards minimal frustration, geometrical constraints, imposed by the native structure, control the folding mechanism and shape the native dynamics revealed by the model. Native-state fluctuations can alternatively be estimated directly from the diversity in the set of NMRstructures for a protein. Based on this information, we identify a highly flexible loop in the ribosomal protein S6 and modify the contact map in a SBM to accommodate the inferred dynamics. By taking into account the probable native state dynamics, the experimental transition state is recovered in the model, and the correct order of folding events is restored. Our study highlights how the shared energy landscape connects folding and function by showing that a better description of the native basin improves the prediction of the folding mechanism
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The physics of bacterial decision making
The choice that bacteria make between sporulation and competence when subjected to stress provides a prototypical example of collective cell fate determination that is stochastic on the individual cell level, yet predictable (deterministic) on the population level. This collective decision is performed by an elaborated gene network. Considerable effort has been devoted to simplify its complexity by taking physics approaches to untangle the basic functional modules that are integrated to form the complete network: (1) A stochastic switch whose transition probability is controlled by two order parametersâpopulation density and internal/external stress. (2) An adaptable timer whose clock rate is normalized by the same two previous order parameters. (3) Sensing units which measure population density and external stress. (4) A communication module that exchanges information about the cells' internal stress levels. (5) An oscillating gate of the stochastic switch which is regulated by the timer. The unique circuit architecture of the gate allows special dynamics and noise management features. The gate opens a window of opportunity in time for competence transitions, during which the circuit generates oscillations that are translated into a chain of short intervals with high transition probability. In addition, the unique architecture of the gate allows filtering of external noise and robustness against variations in circuit parameters and internal noise. We illustrate that a physics approach can be very valuable in investigating the decision process and in identifying its general principles. We also show that both cell-cell variability and noise have important functional roles in the collectively controlled individual decisions
RACIPE: a computational tool for modeling gene regulatory circuits using randomization.
BACKGROUND: One of the major challenges in traditional mathematical modeling of gene regulatory circuits is the insufficient knowledge of kinetic parameters. These parameters are often inferred from existing experimental data and/or educated guesses, which can be time-consuming and error-prone, especially for large networks.
RESULTS: We present a user-friendly computational tool for the community to use our newly developed method named random circuit perturbation (RACIPE), to explore the robust dynamical features of gene regulatory circuits without the requirement of detailed kinetic parameters. Taking the network topology as the only input, RACIPE generates an ensemble of circuit models with distinct randomized parameters and uniquely identifies robust dynamical properties by statistical analysis. Here, we discuss the implementation of the software and the statistical analysis methods of RACIPE-generated data to identify robust gene expression patterns and the functions of genes and regulatory links. Finally, we apply the tool on coupled toggle-switch circuits and a published circuit of B-lymphopoiesis.
CONCLUSIONS: We expect our new computational tool to contribute to a more comprehensive and unbiased understanding of mechanisms underlying gene regulatory networks. RACIPE is a free open source software distributed under (Apache 2.0) license and can be downloaded from GitHub ( https://github.com/simonhb1990/RACIPE-1.0 )
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Redox-dependent gating of VDAC by mitoNEET.
MitoNEET is an outer mitochondrial membrane protein essential for sensing and regulation of iron and reactive oxygen species (ROS) homeostasis. It is a key player in multiple human maladies including diabetes, cancer, neurodegeneration, and Parkinson's diseases. In healthy cells, mitoNEET receives its clusters from the mitochondrion and transfers them to acceptor proteins in a process that could be altered by drugs or during illness. Here, we report that mitoNEET regulates the outer-mitochondrial membrane (OMM) protein voltage-dependent anion channel 1 (VDAC1). VDAC1 is a crucial player in the cross talk between the mitochondria and the cytosol. VDAC proteins function to regulate metabolites, ions, ROS, and fatty acid transport, as well as function as a "governator" sentry for the transport of metabolites and ions between the cytosol and the mitochondria. We find that the redox-sensitive [2Fe-2S] cluster protein mitoNEET gates VDAC1 when mitoNEET is oxidized. Addition of the VDAC inhibitor 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (DIDS) prevents both mitoNEET binding in vitro and mitoNEET-dependent mitochondrial iron accumulation in situ. We find that the DIDS inhibitor does not alter the redox state of MitoNEET. Taken together, our data indicate that mitoNEET regulates VDAC in a redox-dependent manner in cells, closing the pore and likely disrupting VDAC's flow of metabolites
Substrate-Specific Reorganization of the Conformational Ensemble of CSK Implicates Novel Modes of Kinase Function
Protein kinases use ATP as a phosphoryl donor for the posttranslational modification of signaling targets. It is generally
thought that the binding of this nucleotide induces conformational changes leading to closed, more compact forms of the
kinase domain that ideally orient active-site residues for efficient catalysis. The kinase domain is oftentimes flanked by
additional ligand binding domains that up- or down-regulate catalytic function. C-terminal Src kinase (Csk) is a multidomain
tyrosine kinase that is up-regulated by N-terminal SH2 and SH3 domains. Although the X-ray structure of Csk suggests the
enzyme is compact, X-ray scattering studies indicate that the enzyme possesses both compact and open conformational
forms in solution. Here, we investigated whether interactions with the ATP analog AMP-PNP and ADP can shift the
conformational ensemble of Csk in solution using a combination of small angle x-ray scattering and molecular dynamics
simulations. We find that binding of AMP-PNP shifts the ensemble towards more extended rather than more compact
conformations. Binding of ADP further shifts the ensemble towards extended conformations, including highly extended
conformations not adopted by the apo protein, nor by the AMP-PNP bound protein. These ensembles indicate that any
compaction of the kinase domain induced by nucleotide binding does not extend to the overall multi-domain architecture.
Instead, assembly of an ATP-bound kinase domain generates further extended forms of Csk that may have relevance for
kinase scaffolding and Src regulation in the cell
The Unique Cysteine Knot Regulates the Pleotropic Hormone Leptin
Leptin plays a key role in regulating energy intake/expenditure, metabolism and hypertension. It folds into a four-helix
bundle that binds to the extracellular receptor to initiate signaling. Our work on leptin revealed a hidden complexity in the
formation of a previously un-described, cysteine-knotted topology in leptin. We hypothesized that this unique topology
could offer new mechanisms in regulating the protein activity. A combination of in silico simulation and in vitro experiments
was used to probe the role of the knotted topology introduced by the disulphide-bridge on leptin folding and function. Our
results surprisingly show that the free energy landscape is conserved between knotted and unknotted protein, however the
additional complexity added by the knot formation is structurally important. Native state analyses led to the discovery that
the disulphide-bond plays an important role in receptor binding and thus mediate biological activity by local motions on
distal receptor-binding sites, far removed from the disulphide-bridge. Thus, the disulphide-bridge appears to function as a
point of tension that allows dissipation of stress at a distance in leptin
Phylogenetic analysis of eukaryotic NEET proteins uncovers a link between a key gene duplication event and the evolution of vertebrates
NEET proteins belong to a unique family of iron-sulfur proteins in which the 2Fe-2S cluster is coordinated by a CDGSH domain that is followed by the âNEETâ motif. They are involved in the regulation of iron and reactive oxygen metabolism, and have been associated with the progression of diabetes, cancer, aging and neurodegenerative diseases. Despite their important biological functions, the evolution and diversification of eukaryotic NEET proteins are largely unknown. Here we used the three members of the human NEET protein family (CISD1, mitoNEET; CISD2, NAF-1 or Miner 1; and CISD3, Miner2) as our guides to conduct a phylogenetic analysis of eukaryotic NEET proteins and their evolution. Our findings identified the slime mold Dictyostelium discoideumâs CISD proteins as the closest to the ancient archetype of eukaryotic NEET proteins. We further identified CISD3 homologs in fungi that were previously reported not to contain any NEET proteins, and revealed that plants lack homolog(s) of CISD3. Furthermore, our study suggests that the mammalian NEET proteins, mitoNEET (CISD1) and NAF-1 (CISD2), emerged via gene duplication around the origin of vertebrates. Our findings provide new insights into the classification and expansion of the NEET protein family, as well as offer clues to the diverged functions of the human mitoNEET and NAF-1 proteins
Rapid assessment of T-cell receptor specificity of the immune repertoire
Accurate assessment of T-cell-receptor (TCR)âantigen specificity across the whole immune repertoire lies at the heart of improved cancer immunotherapy, but predictive models capable of high-throughput assessment of TCRâpeptide pairs are lacking. Recent advances in deep sequencing and crystallography have enriched the data available for studying TCRâpeptide systems. Here, we introduce RACER, a pairwise energy model capable of rapid assessment of TCRâpeptide affinity for entire immune repertoires. RACER applies supervised machine learning to efficiently and accurately resolve strong TCRâpeptide binding pairs from weak ones. The trained parameters further enable a physical interpretation of interacting patterns encoded in each TCRâpeptide system. When applied to simulate thymic selection of a major-histocompatibility-complex (MHC)-restricted T-cell repertoire, RACER accurately estimates recognition rates for tumor-associated neoantigens and foreign peptides, thus demonstrating its utility in helping address the computational challenge of reliably identifying properties of tumor antigen-specific T-cells at the level of an individual patientâs immune repertoire
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