Understanding how mutations render a drug ineffective is a problem of immense relevance. Often the mechanism through which mutations cause drug resistance can be explained purely through ...thermodynamics. However, the more perplexing situation is when two proteins have the same drug binding affinities but different residence times. In this work, we demonstrate how all‐atom molecular dynamics simulations using recent developments grounded in statistical mechanics can provide a detailed mechanistic rationale for such variances. We discover dissociation mechanisms for the anti‐cancer drug Imatinib (Gleevec) against wild‐type and the N368S mutant of Abl kinase. We show how this point mutation triggers far‐reaching changes in the protein's flexibility and leads to a different, much faster, drug dissociation pathway. We believe that this work marks an efficient and scalable approach to obtain mechanistic insight into resistance mutations in biomolecular receptors that are hard to explain using a structural perspective.
Gleevec has the same binding affinity yet different dissociation times against Wild‐Type and N368S‐mutated Abl kinase. All‐atom enhanced molecular dynamics simulations explain why.
The docking performance of the FRED and HYBRID programs are evaluated on two standardized datasets from the Docking and Scoring Symposium of the ACS Spring 2011 national meeting. The evaluation ...includes cognate docking and virtual screening performance. FRED docks 70 % of the structures to within 2 Å in the cognate docking test. In the virtual screening test, FRED is found to have a mean AUC of 0.75. The HYBRID program uses a modified version of FRED’s algorithm that uses both ligand- and structure-based information to dock molecules, which increases its mean AUC to 0.78. HYBRID can also implicitly account for protein flexibility by making use of multiple crystal structures. Using multiple crystal structures improves HYBRID’s performance (mean AUC 0.80) with a negligible increase in docking time (~15 %).
Membrane contact sites (MCSs) are heterogeneous in shape, composition, and dynamics. Despite this diversity, VAP proteins act as receptors for multiple FFAT motif-containing proteins and drive the ...formation of most MCSs that involve the endoplasmic reticulum (ER). Although the VAP-FFAT interaction is well characterized, no model explains how VAP adapts to its partners in various MCSs. We report that VAP-A localization to different MCSs depends on its intrinsically disordered regions (IDRs) in human cells. VAP-A interaction with PTPIP51 and VPS13A at ER-mitochondria MCS conditions mitochondria fusion by promoting lipid transfer and cardiolipin buildup. VAP-A also enables lipid exchange at ER-Golgi MCS by interacting with oxysterol-binding protein (OSBP) and CERT. However, removing IDRs from VAP-A restricts its distribution and function to ER-mitochondria MCS. Our data suggest that IDRs do not modulate VAP-A preference toward specific partners but do adjust their geometry to MCS organization and lifetime constraints. Thus, IDR-mediated VAP-A conformational flexibility ensures membrane tethering plasticity and efficiency.
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•To examine whether substrate promiscuous enzymes have distinguishing structural features structural attributes of binding/catalytic sites are compared between the curated dataset of ...promiscuous and specialist enzymes (having no known alternate substrate(s)).•Construction of curated datasets of promiscuous and specialist enzymes.•Surprisingly, substrate promiscuous and specialist enzymes have similar binding/catalytic site structural attributes.•The substrate preference of enzymes (promiscuity) could be defined as a continuum feature that varies from narrow to broad range of substrates.
Enzyme promiscuity is the ability of (some) enzymes to perform alternate reactions or catalyze non-cognate substrate(s). The latter is referred to as substrate promiscuity, widely studied for its biotechnological applications and understanding enzyme evolution. Insights into the structural basis of substrate promiscuity would greatly benefit the design and engineering of enzymes. Previous studies on some enzymes have suggested that flexibility, hydrophobicity, and active site protonation state could play an important role in enzyme promiscuity. However, it is not known yet whether substrate promiscuous enzymes have distinctive structural characteristics compared to specialist enzymes, which are specific for a substrate. In pursuit to address this, we have systematically compared substrate/catalytic binding site structural features of substrate promiscuous with those of specialist enzymes. For this, we have carefully constructed dataset of substrate promiscuous and specialist enzymes. On careful analysis, surprisingly, we found that substrate promiscuous and specialist enzymes are similar in various binding/catalytic site structural features such as flexibility, surface area, hydrophobicity, depth, and secondary structures. Recent studies have also alluded that promiscuity is widespread among enzymes. Based on these observations, we propose that substrate promiscuity could be defined as a continuum feature that varies from narrow (specialist) to broad range of substrate preferences. Moreover, diversity of conformational states of an enzyme accessible for ligand binding may possibly regulate its substrate preferences.
The sliding clamp proteins are present in almost all forms of life and participate in various fundamental processes. Many of these proteins accommodate a conserved sequence that interacts with the ...hydrophobic groove on sliding clamps. The conserved sequence on proteins is known as the PCNA-interacting protein box, and the hydrophobic groove of PCNA contains regions of the inter-domain connecting loop, the central loop, and amino acids from the C-terminal tail of PCNA. We performed molecular dynamics simulation studies (1.0 μs) to analyze the structural changes at the atomic level in native, C22Y, and C81R mutant PCNA. Our study revealed significant changes at sites responsible for a functional trimeric form of PCNA. This study also unveils the dynamic behavior of IDCL, central loop, and the C-terminal tail, which are essential regions for protein binding with PCNA and also sheds light on the effect of mutations on binding with the Cdc9 peptide. The observation of Cdc9 peptide complexed with native and mutants (C22Y and C81R) structures possibly reveals the mechanism by which PCNA recruits different proteins required for various biological processes and also highlights the importance of dynamic behavior of key regions involved in PCNA protein-protein interactions.
•MMR deficient mutant PCNA structures compared with native form after binding with yeast Cdc9 peptide.•Key structural changes at the atomic level in native, C22Y, and C81R mutant PCNA upon interaction with Cdc9 peptide.•Specific interactions of PCNA alters the conformational behaviour of interacting regions of the protein.
The translocator protein (TSPO), an 18-kDa transmembrane protein primarily found in the outer mitochondrial membrane, is evolutionarily conserved and widely distributed across species. In mammals, ...TSPO has been described as a key member of a multiprotein complex involved in many putative functions and, over the years, several classes of ligand have been developed to modulate these functions. In this review, we consider the currently available atomic structures of mouse and bacterial TSPO and propose a rationale for the development of new ligands for the protein. We provide a review of TSPO monomeric and oligomeric states and their conformational flexibility, together with ligand-binding site and interaction mechanisms. These data are expected to help considerably the development of high-affinity ligands for TSPO-based therapies or diagnostics.
The transmembrane protein TSPO has been described as a diagnostic and therapeutic target for inflammation and, in particular, brain diseases.Various classes of TSPO ligand have been developed since its discovery in 1977, and some are routinely used for diagnosis and treatment.The design of new ligands is needed not only to improve cellular specificity, for instance to distinguish abnormal cells, but also for targeting to TSPO as part of a multiprotein complex of varying composition depending on the cell.Drug design will gain from TSPO structure–function analysis and computer simulation methods, such as molecular dynamics.
Structure-based virtual screening is currently an established tool in drug lead discovery projects. Although in the last years the field saw an impressive progress in terms of algorithm development, ...computational performance, and retrospective and prospective applications in ligand identification, there are still long-standing challenges where further improvement is needed. In this review, we consider the conceptual frame, state-of-the-art and recent developments of three critical “structural” issues in structure-based drug lead discovery: the use of homology modeling to accurately model the binding site when no experimental structures are available, the necessity of accounting for the dynamics of intrinsically flexible systems as proteins, and the importance of considering active site water molecules in lead identification and optimization campaigns.
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•Structure-based virtual screening in drug lead discovery and design.•We review the conceptual frame, recent developments and standing challenges.•Homology modeling can furnish accurate binding site models for virtual screening.•Intrinsic target dynamics consideration enhances the outcome of screening campaigns.•Algorithms to rationalize active site water molecules are increasingly used.
Copper is an essential element in nature but in excess, it is toxic to the living cell. The human metallochaperone Atox1 participates in copper homeostasis and is responsible for copper transmission. ...In a previous multiscale simulation study, we noticed a change in the coordination state of the Cu(I) ion, from 4 bound cysteine residues to 3, in agreement with earlier studies. Here, we perform and analyze classical molecular dynamic simulations of various coordination states: 2, 3, and 4. The main observation is an increase in protein flexibility as a result of a decrease in the coordination state. In addition, we identified several populated conformations that correlate well with double electron–electron resonance distance distributions or an X‐ray structure of Cu(I)‐bound Atox1. We suggest that the increased flexibility might benefit the process of ion transmission between interacting proteins. Further experiments can scrutinize this hypothesis and shed additional light on the mechanism of action of Atox1.
•Connector regions were found to have structural variation between pepsin and renin.•pH stability may be defined by altered amino acid composition in connector regions.•The N-terminus of pepsin is ...more flexible than the N-terminus renin.•Pepsin may unfold in one event; renin may unfold in a series of smaller events.•Pepsin has more predicted regions that may initiate unfolding compared to renin.
Pepsin, the archetypal pepsin-like aspartic protease, is irreversibly denatured when exposed to neutral pH conditions whereas renin, a structural homologue of pepsin, is fully stable and optimally active in the same conditions despite sharing highly similar enzyme architecture. To gain insight into the structural determinants of differential aspartic protease pH stability, the present study used comparative bioinformatic and structural analyses. In pepsin, an abundance of polar and aspartic acid residues were identified, a common trait with other acid-stable enzymes. Conversely, renin was shown to have increased levels of basic amino acids. In both pepsin and renin, the solvent exposure of these charged groups was high. Having similar overall acidic residue content, the solvent-exposed basic residues may allow for extensive salt bridge formation in renin, whereas in pepsin, these residues are protonated and serve to form stabilizing hydrogen bonds at low pH. Relative differences in structure and sequence in the turn and joint regions of the β-barrel and ψ-loop in both the N- and C-terminal lobes were identified as regions of interest in defining divergent pH stability. Compared to the structural rigidity of renin, pepsin has more instability associated with the N-terminus, specifically the B/C connector. By contrast, renin exhibits greater C-terminal instability in turn and connector regions. Overall, flexibility differences in connector regions, and amino acid composition, particularly in turn and joint regions of the β-barrel and ψ-loops, likely play defining roles in determining pH stability for renin and pepsin.
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