Browsing by Author "Lammert, Heiko"

Now showing 1 - 8 of 8
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    A magnesium-induced triplex pre-organizes the SAM-II riboswitch
    (Public Library of Science, 2017) Roy, Susmita; Lammert, Heiko; Hayes, Ryan L.; Chen, Bin; LeBlanc, Regan; Dayie, T.Kwaku; Onuchic, José Nelson; Sanbonmatsu, Karissa Y.; Center for Theoretical Biological Physics
    Our 13C- and 1H-chemical exchange saturation transfer (CEST) experiments previously revealed a dynamic exchange between partially closed and open conformations of the SAM-II riboswitch in the absence of ligand. Here, all-atom structure-based molecular simulations, with the electrostatic effects of Manning counter-ion condensation and explicit magnesium ions are employed to calculate the folding free energy landscape of the SAM-II riboswitch. We use this analysis to predict that magnesium ions remodel the landscape, shifting the equilibrium away from the extended, partially unfolded state towards a compact, pre-organized conformation that resembles the ligand-bound state. Our CEST and SAXS experiments, at different magnesium ion concentrations, quantitatively confirm our simulation results, demonstrating that magnesium ions induce collapse and pre-organization. Agreement between theory and experiment bolsters microscopic interpretation of our simulations, which shows that triplex formation between helix P2b and loop L1 is highly sensitive to magnesium and plays a key role in pre-organization. Pre-organization of the SAM-II riboswitch allows rapid detection of ligand with high selectivity, which is important for biological function.
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    Altered Backbone and Side-Chain Interactions Result in Route Heterogeneity during the Folding of Interleukin-1b (IL-1b)
    (Biophysical Society, 2013-08) Capraro, Dominique T.; Lammert, Heiko; Heidary, David K.; Roy, Melinda; Gross, Larry A.; Onuchic, José N.; Jennings, Patricia A.; Center for Theoretical Biological Physics
    Deletion of the b-bulge trigger-loop results in both a switch in the preferred folding route, from the functional loop packing folding route to barrel closure, as well as conversion of the agonist activity of IL-1b into antagonist activity. Conversely, circular permutations of IL-1b conserve the functional folding route as well as the agonist activity. These two extremes in the folding-functional interplay beg the question of whether mutations in IL-1b would result in changes in the populations of heterogeneous folding routes and the signaling activity. A series of topologically equivalent water-mediated b-strand bridging interactions within the pseudosymmetric b-trefoil fold of IL-1b highlight the backbone water interactions that stabilize the secondary and tertiary structure of the protein. Additionally, conserved aromatic residues lining the central cavity appear to be essential for both stability and folding. Here, we probe these protein backbone-water molecule and side chain-side chain interactions and the role they play in the folding mechanism of this geometrically stressed molecule. We used folding simulations with structure-based models, as well as a series of folding kinetic experiments to examine the effects of the F42W core mutation on the folding landscape of IL-1b. This mutation alters water-mediated backbone interactions essential for maintaining the trefoil fold. Our results clearly indicate that this perturbation in the primary structure alters a structural water interaction and consequently modulates the population of folding routes accessed during folding and signaling activity.
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    Constructing a folding model for protein S6 guided by native fluctuations deduced from NMR structures
    (AIP Publishing LLC., 2015) Lammert, Heiko; Noel, Jeffrey K.; Haglund, Ellinor; Schug, Alexander; Onuchic, José N.; Center for Theoretical Biological Physics
    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 protein’s 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 Dominant Folding Route Minimizes Backbone Distortion in SH3
    (2012-11-15) Lammert, Heiko; Noel, Jeffrey K.; Onuchic, José N.; Center for Theoretical Biological Physics
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    Magnesium controls aptamer-expression platform switching in the SAM-I riboswitch
    (Oxford University Press, 2019) Roy, Susmita; Hennelly, Scott P.; Lammert, Heiko; Onuchic, José Nelson; Sanbonmatsu, Karissa Y.
    Investigations of most riboswitches remain confined to the ligand-binding aptamer domain. However, during the riboswitch mediated transcription regulation process, the aptamer domain and the expression platform compete for a shared strand. If the expression platform dominates, an anti-terminator helix is formed, and the transcription process is active (ON state). When the aptamer dominates, transcription is terminated (OFF state). Here, we use an expression platform switching experimental assay and structure-based electrostatic simulations to investigate this ON-OFF transition of the full length SAM-I riboswitch and its magnesium concentration dependence. Interestingly, we find the ratio of the OFF population to the ON population to vary non-monotonically as magnesium concentration increases. Upon addition of magnesium, the aptamer domain pre-organizes, populating the OFF state, but only up to an intermediate magnesium concentration level. Higher magnesium concentration preferentially stabilizes the anti-terminator helix, populating the ON state, relatively destabilizing the OFF state. Magnesium mediated aptamer-expression platform domain closure explains this relative destabilization of the OFF state at higher magnesium concentration. Our study reveals the functional potential of magnesium in controlling transcription of its downstream genes and underscores the importance of a narrow concentration regime near the physiological magnesium concentration ranges, striking a balance between the OFF and ON states in bacterial gene regulation.
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    Pierced Lasso Bundles Are a New Class of Knot-like Motifs
    (Public Library of Science, 2014) Haglund, Ellinor; Sulkowska, Joanna I.; Noel, Jeffrey K.; Lammert, Heiko; Onuchic, José Nelson; Jennings, Patricia A.; Center for Theoretical Biological Physics
    A four-helix bundle is a well-characterized motif often used as a target for designed pharmaceutical therapeutics and nutritional supplements. Recently, we discovered a new structural complexity within this motif created by a disulphide bridge in the long-chain helical bundle cytokine epileptic When oxidized, leptin contains a disulphide bridge creating a covalent-loop through which part of the polypeptide chain is threaded (as seen in knotted proteins). We explored whether other proteins contain a similar intriguing knot-like structure as in leptin and discovered 11 structurally homologous proteins in the PDB. We call this new helical family class the Pierced Lasso Bundle (PLB) and the knot-like threaded structural motif a Pierced Lasso (PL). In the current study, we use structure-based simulation to investigate the threading/folding mechanisms for all the PLBs along with three unthreaded homologs as the covalent loop (or lasso) in leptin is important in folding dynamics and activity. We find that the presence of a small covalent loop leads to a mechanism where structural elements slipknot to thread through the covalent loop. Larger loops use a piercing mechanism where the free terminal plugs through the covalent loop. Remarkably, the position of the loop as well as its size influences the native state dynamics, which can impact receptor binding and biological activity. This previously unrecognized complexity of knot-like proteins within the helical bundle family comprises a completely new class within the knot family, and the hidden complexity we unraveled in the PLBs is expected to be found in other protein structures outside the four-helix bundles. The insights gained here provide critical new elements for future investigation of this emerging class of proteins, where function and the energetic landscape can be controlled by hidden topology, and should be take into account in ab initio predictions of newly identified protein targets.
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    SMOG 2: A Versatile Software Package for Generating Structure-Based Models
    (Public Library of Science, 2016) Noel, Jeffrey K.; Levi, Mariana; Raghunathan, Mohit; Lammert, Heiko; Hayes, Ryan L.; Onuchic, José N.; Whitford, Paul C.; Center for Theoretical Biological Physics
    Molecular dynamics simulations with coarse-grained or simplified Hamiltonians have proven to be an effective means of capturing the functionally important long-time and large-length scale motions of proteins and RNAs. Originally developed in the context of protein folding, structure-based models (SBMs) have since been extended to probe a diverse range of biomolecular processes, spanning from protein and RNA folding to functional transitions in molecular machines. The hallmark feature of a structure-based model is that part, or all, of the potential energy function is defined by a known structure. Within this general class of models, there exist many possible variations in resolution and energetic composition. SMOG 2 is a downloadable software package that reads user-designated structural information and user-defined energy definitions, in order to produce the files necessary to use SBMs with high performance molecular dynamics packages: GROMACS and NAMD. SMOG 2 is bundled with XML-formatted template files that define commonly used SBMs, and it can process template files that are altered according to the needs of each user. This computational infrastructure also allows for experimental or bioinformatics-derived restraints or novel structural features to be included, e.g. novel ligands, prosthetic groups and post-translational/transcriptional modifications. The code and user guide can be downloaded at http://smog-server.org/smog2.
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    The Dominant Folding Route Minimizes Backbone Distortion in SH3
    (Public Library of Science, 2012) Lammert, Heiko; Noel, Jeffrey K.; Onuchic, José N.; Center for Theoretical Biological Physics
    Energetic frustration in protein folding is minimized by evolution to create a smooth and robust energy landscape. As a result the geometry of the native structure provides key constraints that shape protein folding mechanisms. Chain connectivity in particular has been identified as an essential component for realistic behavior of protein folding models. We study the quantitative balance of energetic and geometrical influences on the folding of SH3 in a structure-based model with minimal energetic frustration. A decomposition of the two-dimensional free energy landscape for the folding reaction into relevant energy and entropy contributions reveals that the entropy of the chain is not responsible for the folding mechanism. Instead the preferred folding route through the transition state arises from a cooperative energetic effect. Off-pathway structures are penalized by excess distortion in local backbone configurations and contact pair distances. This energy cost is a new ingredient in the malleable balance of interactions that controls the choice of routes during protein folding.