RESUMO
We modeled 3D structures of all SARS-CoV-2 proteins, generating 2,060 models that span 69% of the viral proteome and provide details not available elsewhere. We found that Ë6% of the proteome mimicked human proteins, while Ë7% was implicated in hijacking mechanisms that reverse post-translational modifications, block host translation, and disable host defenses; a further Ë29% self-assembled into heteromeric states that provided insight into how the viral replication and translation complex forms. To make these 3D models more accessible, we devised a structural coverage map, a novel visualization method to show what is-and is not-known about the 3D structure of the viral proteome. We integrated the coverage map into an accompanying online resource (https://aquaria.ws/covid) that can be used to find and explore models corresponding to the 79 structural states identified in this work. The resulting Aquaria-COVID resource helps scientists use emerging structural data to understand the mechanisms underlying coronavirus infection and draws attention to the 31% of the viral proteome that remains structurally unknown or dark.
Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , Interações Hospedeiro-Patógeno/genética , Processamento de Proteína Pós-Traducional , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/química , Sistemas de Transporte de Aminoácidos Neutros/genética , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Enzima de Conversão de Angiotensina 2/química , Enzima de Conversão de Angiotensina 2/genética , Sítios de Ligação , COVID-19/genética , COVID-19/metabolismo , COVID-19/virologia , Biologia Computacional/métodos , Proteínas do Envelope de Coronavírus/química , Proteínas do Envelope de Coronavírus/genética , Proteínas do Envelope de Coronavírus/metabolismo , Proteínas do Nucleocapsídeo de Coronavírus/química , Proteínas do Nucleocapsídeo de Coronavírus/genética , Proteínas do Nucleocapsídeo de Coronavírus/metabolismo , Humanos , Proteínas de Transporte da Membrana Mitocondrial/química , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Modelos Moleculares , Mimetismo Molecular , Neuropilina-1/química , Neuropilina-1/genética , Neuropilina-1/metabolismo , Fosfoproteínas/química , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas/métodos , Multimerização Proteica , SARS-CoV-2/química , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Proteínas da Matriz Viral/química , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/metabolismo , Proteínas Viroporinas/química , Proteínas Viroporinas/genética , Proteínas Viroporinas/metabolismo , Replicação ViralRESUMO
Human leucocyte antigen (HLA) class II molecules in humans are encoded by three different loci, HLA-DR, -DQ, and -DP. These molecules share approximately 70% sequence similarity and all present peptide ligands to circulating T cells. While the peptide repertoires of numerous HLA-DR, -DQ, and -DP allotypes have been examined, there have been few reports on the combined repertoire of these co-inherited molecules expressed in a single cell as an extended HLA haplotype. Here we describe the endogenous peptide repertoire of a human B lymphoblastoid cell line (C1R) expressing the class II haplotype HLA-DR12/DQ7/DP4. We have identified 71350 unique naturally processed peptides presented collectively by HLA-DR12, HLA-DQ7, or HLA-DP4. The resulting "haplodome" is complemented by the cellular proteome defined by standard LC-MS/MS approaches. This large dataset has shed light on properties of these class II ligands especially the preference for membrane and extracellular source proteins. Our data also provides insights into the co-evolution of these conserved haplotypes of closely linked and co-inherited HLA molecules; which together increase sequence coverage of cellular proteins for immune surveillance with minimal overlap between each co-inherited HLA-class II allomorph.
Assuntos
Antígenos HLA-DQ , Proteoma , Cromatografia Líquida , Antígenos HLA-DQ/genética , Haplótipos , Antígenos de Histocompatibilidade Classe II , Humanos , Monitorização Imunológica , Espectrometria de Massas em TandemRESUMO
Protein conformational changes and dynamic behavior are fundamental for such processes as catalysis, regulation, and substrate recognition. Although protein dynamics have been successfully explored in computer simulation, there is an intermediate-scale of motions that has proven difficult to simulate - the motion of individual segments or domains that move independently of the body the protein. Here, we introduce a molecular-dynamics perturbation method, the Rotamerically Induced Perturbation (RIP), which can generate large, coherent motions of structural elements in picoseconds by applying large torsional perturbations to individual sidechains. Despite the large-scale motions, secondary structure elements remain intact without the need for applying backbone positional restraints. Owing to its computational efficiency, RIP can be applied to every residue in a protein, producing a global map of deformability. This map is remarkably sparse, with the dominant sites of deformation generally found on the protein surface. The global map can be used to identify loops and helices that are less tightly bound to the protein and thus are likely sites of dynamic modulation that may have important functional consequences. Additionally, they identify individual residues that have the potential to drive large-scale coherent conformational change. Applying RIP to two well-studied proteins, Dihdydrofolate Reductase and Triosephosphate Isomerase, which possess functionally-relevant mobile loops that fluctuate on the microsecond/millisecond timescale, the RIP deformation map identifies and recapitulates the flexibility of these elements. In contrast, the RIP deformation map of alpha-lytic protease, a kinetically stable protein, results in a map with no significant deformations. In the N-terminal domain of HSP90, the RIP deformation map clearly identifies the ligand-binding lid as a highly flexible region capable of large conformational changes. In the Estrogen Receptor ligand-binding domain, the RIP deformation map is quite sparse except for one large conformational change involving Helix-12, which is the structural element that allosterically links ligand binding to receptor activation. RIP analysis has the potential to discover sites of functional conformational changes and the linchpin residues critical in determining these conformational states.
Assuntos
Modelos Químicos , Modelos Moleculares , Análise de Sequência de Proteína/métodos , Triose-Fosfato Isomerase/química , Triose-Fosfato Isomerase/ultraestrutura , Sequência de Aminoácidos , Simulação por Computador , Dados de Sequência Molecular , Conformação ProteicaRESUMO
BACKGROUND: An accurate rendering of interior surfaces can facilitate the analysis of mechanisms at atomic-level detail, such as the transport of substrates in the ammonia channel. In molecular viewers, one must remove the exterior surface that obscures the channel surface by clipping the viewing plane or manually selecting the channel residues in order to display a partial surface. Neither method is entirely satisfactory, as unwanted additional pieces of surfaces are always generated. RESULTS: To cleanly visualize a channel surface, we present HOLLOW, a program that generates a "casting" of the interior volume of the protein as dummy atoms. We show that the molecular surface of the dummy atoms closely approximates the channel surface, where this complementary surface of the protein channel can be displayed without superfluous surfaces. CONCLUSION: The use of HOLLOW significantly simplifies the generation of channel surfaces, and other interior surfaces of protein structures. HOLLOW is written in PYTHON and is available at http://hollow.sourceforge.net.
Assuntos
Modelos Moleculares , Software , Gráficos por Computador , Estrutura Molecular , Conformação Proteica , Propriedades de SuperfícieRESUMO
BACKGROUND: An important element in homology modeling is the use of rotamers to parameterize the sidechain conformation. Despite the many libraries of sidechain rotamers that have been developed, a number of rotamers have been overlooked, due to the fact that they involve hydrogen atoms. RESULTS: We identify new, well-populated rotamers that involve the hydroxyl-hydrogen atoms of Ser, Thr and Tyr, and the sulfhydryl-hydrogen atom of Cys, using high-resolution crystal structures (<1.2 A). Although there were refinement artifacts in these structures, comparison with the electron-density maps allowed the placement of hydrogen atoms involved in hydrogen bonds. The chi2 rotamers in Ser, Thr and Cys are consistent with tetrahedral bonding, while the chi3 rotamers in Tyr are consistent with trigonal-planar bonding. Similar rotamers are found in hydrogen atoms that were computationally placed with the Reduce program from the Richardson lab. CONCLUSION: Knowledge of these new rotamers will improve the evaluation of hydrogen-bonding networks in protein structures.
Assuntos
Aminoácidos/química , Hidrogênio/química , Radical Hidroxila/química , Conformação Proteica , Compostos de Sulfidrila/química , Cristalografia por Raios X , Cisteína/química , Ligação de Hidrogênio , Estrutura Secundária de Proteína , Serina/química , Treonina/química , Tirosina/químicaRESUMO
Peptides often have conformational preferences. We simulated 133 peptide 8-mer fragments from six different proteins, sampled by replica-exchange molecular dynamics using Amber7 with a GB/SA (generalized-Born/solvent-accessible electrostatic approximation to water) implicit solvent. We found that 85 of the peptides have no preferred structure, while 48 of them converge to a preferred structure. In 85% of the converged cases (41 peptides), the structures found by the simulations bear some resemblance to their native structures, based on a coarse-grained backbone description. In particular, all seven of the beta hairpins in the native structures contain a fragment in the turn that is highly structured. In the eight cases where the bioinformatics-based I-sites library picks out native-like structures, the present simulations are largely in agreement. Such physics-based modeling may be useful for identifying early nuclei in folding kinetics and for assisting in protein-structure prediction methods that utilize the assembly of peptide fragments.
Assuntos
Dobramento de Proteína , Fenômenos Biofísicos , Biofísica , Biologia Computacional/métodos , Simulação por Computador , Entropia , Cinética , Modelos Moleculares , Modelos Estatísticos , Conformação Molecular , Peptídeos/química , Conformação Proteica , Estrutura Secundária de Proteína , Proteômica/métodos , Solventes/químicaRESUMO
Major histocompatibility complex class I (MHC-I) molecules play a crucial role in immunity by capturing peptides for presentation to T cells and natural killer (NK) cells. The peptide termini are tethered within the MHC-I antigen-binding groove, but it is unknown whether other presentation modes occur. Here we show that 20% of the HLA-B*57:01 peptide repertoire comprises N-terminally extended sets characterized by a common motif at position 1 (P1) to P2. Structures of HLA-B*57:01 presenting N-terminally extended peptides, including the immunodominant HIV-1 Gag epitope TW10 (TSTLQEQIGW), showed that the N terminus protrudes from the peptide-binding groove. The common escape mutant TSNLQEQIGW bound HLA-B*57:01 canonically, adopting a dramatically different conformation than the TW10 peptide. This affected recognition by killer cell immunoglobulin-like receptor (KIR) 3DL1 expressed on NK cells. We thus define a previously uncharacterized feature of the human leukocyte antigen class I (HLA-I) immunopeptidome that has implications for viral immune escape. We further suggest that recognition of the HLA-B*57:01-TW10 epitope is governed by a 'molecular tension' between the adaptive and innate immune systems.
Assuntos
HIV-1/metabolismo , Antígenos de Histocompatibilidade Classe I/química , Evasão da Resposta Imune , Peptídeos/química , Sequência de Aminoácidos , Cristalografia por Raios X , Epitopos/química , Células HEK293 , Humanos , Metaboloma , Proteínas Mutantes/química , Mutação/genética , Peptídeos/metabolismo , Ligação Proteica , Estrutura Quaternária de Proteína , Receptores KIR3DL1/metabolismo , Ressonância de Plasmônio de Superfície , Produtos do Gene gag do Vírus da Imunodeficiência Humana/química , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismoRESUMO
In proteins, the proline ring exists predominantly in two discrete states. However, there is also a small but significant amount of flexibility in the proline ring of high-resolution protein structures. We have found that this side-chain flexibility is coupled to the backbone conformation. To study this coupling, we have developed a model that is simply based on geometric and steric factors and not on energetics. We show that the coupling between phi and chi1 torsions in the proline ring can be described by an analytic equation that was developed by Bricard in 1897, and we describe a computer algorithm that implements the equation. The model predicts the observed coupling very well. The strain in the C(gamma)-C(delta)-N angle appears to be the principal barrier between the UP and DOWN pucker. This strain is relaxed to allow the proline ring to flatten in the rare PLANAR conformation.
Assuntos
Prolina/química , Proteínas/química , Algoritmos , Modelos Moleculares , Estrutura MolecularRESUMO
The structures of the beta-sheets and the beta-ribbons have been analysed using high-resolution protein structure data. Systematic asymmetries measured in both parallel and antiparallel beta-structures include the sheet twist and the strand shear. In order to determine the origin of these asymmetries, numerous interactions and correlations were examined. The strongest correlations are observed for residues in antiparallel beta-sheets and beta-ribbons that form non-H-bonded pairs. For these residues, the sheet twist is correlated to the backbone phi angle but not to the psi angle. Our analysis supports the existence of an inter-strand C(alpha)H(alpha)...O weak H-bond, which, together with the CO...HN H-bond, constitutes a bifurcated H-bond that links neighbouring beta-strands. Residues of beta-sheets and beta-ribbons in high-resolution protein structures form a distinct region of the Ramachandran plot, which is determined by the formation of the bifurcated H-bond, the formation of an intra-strand O...H(alpha) non-bonded polar interaction, and an intra-strand O...C(beta) steric clash. Using beta-strands parameterised by phi-psi values from the allowed beta-sheet region of the Ramachandran plot, the shear and the right-hand twist can be reproduced in a simple model of the antiparallel and parallel beta-ribbon that models the bifurcated H-bonds specifically. The conformations of interior residues of beta-sheets are shown to be subsets of the conformations of residues of beta-ribbons.
Assuntos
Modelos Moleculares , Estrutura Secundária de Proteína , Proteínas/química , Bases de Dados de Proteínas , Ligação de HidrogênioRESUMO
BACKGROUND: The Ramachandran plot is a fundamental tool in the analysis of protein structures. Of the 4 basic types of Ramachandran plots, the interactions that determine the generic and proline Ramachandran plots are well understood. The interactions of the glycine and pre-proline Ramachandran plots are not. RESULTS: In glycine, the psi angle is typically clustered at psi = 180 degrees and psi = 0 degrees. We show that these clusters correspond to conformations where either the N(i+1) or O atom is sandwiched between the two Halpha atoms of glycine. We show that the shape of the 5 distinct regions of density (the alpha, alphaL, betaS, betaP and betaPR regions) can be reproduced with electrostatic dipole-dipole interactions. In pre-proline, we analyse the origin of the zeta region of the Ramachandran plot, a region unique to pre-proline. We show that it is stabilized by a CO(i-1)...CdeltaHdelta(i+1) weak hydrogen bond. This is analogous to the CO(i-1)...NH(i+1) hydrogen bond that stabilizes the gamma region in the generic Ramachandran plot. CONCLUSION: We have identified the specific interactions that affect the backbone of glycine and pre-proline. Knowledge of these interactions will improve current force-fields, and help understand structural motifs containing these residues.
Assuntos
Biologia Computacional/métodos , Glicina/química , Prolina/química , Proteínas/química , Motivos de Aminoácidos , Fenômenos Biofísicos , Biofísica , Carbono/química , Bases de Dados de Proteínas , Ligação de Hidrogênio , Ligação Proteica , Conformação Proteica , Software , Eletricidade Estática , EstereoisomerismoRESUMO
What determines the shape of the allowed regions in the Ramachandran plot? Although Ramachandran explained these regions in terms of 1-4 hard-sphere repulsions, there are discrepancies with the data where, in particular, the alphaR, alphaL, and beta-strand regions are diagonal. The alphaR-region also varies along the alpha-helix where it is constrained at the center and the amino terminus but diffuse at the carboxyl terminus. By analyzing a high-resolution database of protein structures, we find that certain 1-4 hard-sphere repulsions in the standard steric map of Ramachandran do not affect the statistical distributions. By ignoring these steric clashes (NH(i+1) and O(i-1)C), we identify a revised set of steric clashes (CbetaO, O(i-1)N(i+1), CbetaN(i+1), O(i-1)Cbeta, and O(i-1)O) that produce a better match with the data. We also find that the strictly forbidden region in the Ramachandran plot is excluded by multiple steric clashes, whereas the outlier region is excluded by only one significant steric clash. However, steric clashes alone do not account for the diagonal regions. Using electrostatics to analyze the conformational dependence of specific interatomic interactions, we find that the diagonal shape of the alphaR and alphaL-regions also depends on the optimization of the NH(i+1) and O(i-1)C interactions, and the diagonal beta-strand region is due to the alignment of the CO and NH dipoles. Finally, we reproduce the variation of the Ramachandran plot along the alpha-helix in a simple model that uses only H-bonding constraints. This allows us to rationalize the difference between the amino terminus and the carboxyl terminus of the alpha-helix in terms of backbone entropy.
Assuntos
Estrutura Secundária de Proteína , Proteínas/química , Aminoácidos/química , Ligação de Hidrogênio , Modelos Moleculares , Conformação Molecular , Eletricidade EstáticaRESUMO
BACKGROUND: The annotation of surface exposed bacterial membrane proteins is an important step in interpretation and validation of proteomic experiments. In particular, proteins detected by cell surface protease shaving experiments can indicate exposed regions of membrane proteins that may contain antigenic determinants or constitute vaccine targets in pathogenic bacteria. RESULTS: Inmembrane is a tool to predict the membrane proteins with surface-exposed regions of polypeptide in sets of bacterial protein sequences. We have re-implemented a protocol for Gram-positive bacterial proteomes, and developed a new protocol for Gram-negative bacteria, which interface with multiple predictors of subcellular localization and membrane protein topology. Through the use of a modern scripting language, inmembrane provides an accessible code-base and extensible architecture that is amenable to modification for related sequence annotation tasks. CONCLUSIONS: Inmembrane easily integrates predictions from both local binaries and web-based queries to help gain an overview of likely surface exposed protein in a bacterial proteome. The program is hosted on the Github repository http://github.com/boscoh/inmembrane.
RESUMO
In structural biology, management of a large number of Protein Data Bank (PDB) files and raw X-ray diffraction images often presents a major organizational problem. Existing software packages that manipulate these file types were not designed for these kinds of file-management tasks. This is typically encountered when browsing through a folder of hundreds of X-ray images, with the aim of rapidly inspecting the diffraction quality of a data set. To solve this problem, a useful functionality of the Macintosh operating system (OSX) has been exploited that allows custom visualization plugins to be attached to certain file types. Software plugins have been developed for diffraction images and PDB files, which in many scenarios can save considerable time and effort. The direct visualization of diffraction images and PDB structures in the file browser can be used to identify key files of interest simply by scrolling through a list of files.
RESUMO
Molecular dynamics (MD) simulation is an established method for studying the conformational changes that are important for protein function. Recent advances in hardware and software have allowed MD simulations over the same timescales as experiment, improving the agreement between theory and experiment to a large extent. However, running such simulations are costly, in terms of resources, storage, and trajectory analysis. There is still a place for techniques that involve short MD simulations. In order to overcome the sampling paucity of short time-scales, hybrid methods that include some form of MD simulation can exploit certain features of the system of interest, often combining experimental information in surprising ways. Here, we review some recent hybrid approaches to the simulation of proteins.
Assuntos
Simulação de Dinâmica Molecular , Proteínas/química , Ligantes , Ligação Proteica , Conformação Proteica , Proteínas/metabolismo , Transdução de SinaisRESUMO
Single-domain allostery has been postulated to occur through intramolecular pathways of signaling within a protein structure. We had previously investigated these pathways by introducing a local thermal perturbation and analyzed the anisotropic propagation of structural changes throughout the protein. Here, we develop an improved approach, the Rotamerically Induced Perturbation (RIP), that identifies strong couplings between residues by analyzing the pathways of heat-flow resulting from thermal excitation of rotameric rotations at individual residues. To explore the nature of these couplings, we calculate the complete coupling maps of 5 different PDZ domains. Although the PDZ domain is a well conserved structural fold that serves as a scaffold in many protein-protein complexes, different PDZ domains display unique patterns of conformational flexibility in response to ligand binding: some show a significant shift in a set of alpha-helices, while others do not. Analysis of the coupling maps suggests a simple relationship between the computed couplings and observed conformational flexibility. In domains where the alpha-helices are rigid, we find couplings of the alpha-helices to the body of the protein, whereas in domains having ligand-responsive alpha-helices, no couplings are found. This leads to a model where the alpha-helices are intrinsically dynamic but can be damped if sidechains interact at key tertiary contacts. These tertiary contacts correlate to high covariation contacts as identified by the statistical coupling analysis method. As these dynamic modules are exploited by various allosteric mechanisms, these tertiary contacts have been conserved by evolution.
Assuntos
Sequência Conservada , Domínios PDZ , Sequência de Aminoácidos , Temperatura Alta , Computação Matemática , Dobramento de Proteína , Estrutura Secundária de Proteína , RotaçãoRESUMO
One of the applications of Molecular Dynamics (MD) simulations is to explore the energetic barriers to mechanical unfolding of proteins such as occurs in response to the mechanical pulling of single molecules in Atomic Force Microscopy (AFM) experiments. Although Steered Molecular Dynamics simulations have provided microscopic details of the unfolding process during the pulling, the simulated forces required for unfolding are typically far in excess of the measured values. To rectify this, we have developed the Pulsed Unconstrained Fluctuating Forces (PUFF) method, which induces constant-momentum motions by applying forces directly to the instantaneous velocity of selected atoms in a protein system. The driving forces are applied in pulses, which allows the system to relax between pulses, resulting in more accurate unfolding force estimations than in previous methods. In the cases of titin, ubiquitin and e2lip3, the PUFF trajectories produce force fluctuations that agree quantitatively with AFM experiments. Another useful property of PUFF is that simulations get trapped if the target momentum is too low, simplifying the discovery and analysis of unfolding intermediates.