Roger Armen - Academia.edu (original) (raw)

Papers by Roger Armen

Circulation, 2014

Introduction: The Akt kinase isoforms (Akt1, Akt2 and Akt3) are activated downstream of the insul... more Introduction: The Akt kinase isoforms (Akt1, Akt2 and Akt3) are activated downstream of the insulin receptor, and exert unique effects on cell growth, survival and metabolism. We recently described a novel ATP-dependent "dephosphorylation-resistance cage" in Akt kinases that controls access of cellular phosphatases to dephosphorylate the Akt activation loop (T308 in Akt1). Hypothesis: Here, we describe that, when Akt is localized at the cell membrane, intramolecular interactions of Akt C-terminal sequences with the hydrophobic groove of the kinase domain protect the phosphorylated activation loop (pT308 in Akt1) from cellular phosphatases. Methods and Results: Charged amino acid replacements of Akt C-terminal phosphorylation sites regulated by MTORC2 (Ser473) and by cyclin-dependent kinase 2 (Ser477) increased phospho-T308 protection from cellular phosphatases regardless of subcellular location. Functionally, these phosphatase-resistant Akt variants were refractory to cera...

Journal of Computational Chemistry, 2017

Following insights from recent crystal structures of the muscarinic acetylcholine receptor, bindi... more Following insights from recent crystal structures of the muscarinic acetylcholine receptor, binding modes of Positive Allosteric Modulators (PAMs) were predicted under the assumption that PAMs should bind to the extracellular surface of the active state. A series of well-characterized PAMs for adenosine (A 1 R, A 2A R, A 3 R) and muscarinic acetylcholine (M 1 R, M 5 R) receptors were modeled using both rigid and flexible receptor CHARMM-based molecular docking. Studies of adenosine receptors investigated the molecular basis of the probe-dependence of PAM activity by modeling in complex with specific agonist radioligands. Consensus binding modes map common pharmacophore features of several chemical series to specific binding interactions. These models provide a rationalization of how PAM binding slows agonist radioligand dissociation kinetics. M 1 R PAMs were predicted to bind in the analogous M 2 R PAM LY2119620 binding site. The M 5 R NAM (ML-375) was predicted to bind in the PAM (ML-380) binding site with a unique inducedfit receptor conformation.

The Biochemical journal, Jan 22, 2015

The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signal... more The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signaling and regulate cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (T308 in Akt1) in an ATP-dependent manner. In this report, we describe a distinct mechanism to control T308 dephosphorylation, and thus Akt deactivation, that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal MTORC2 phosphorylation site (Ser473) increased phosphatase resistance of the phosphorylated activation loop (pT308) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent T308 phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydropho...

We present a scalable and accurate method for classifying protein-ligand binding geometries in mo... more We present a scalable and accurate method for classifying protein-ligand binding geometries in molecular docking. Our method is a three-step process: the first step encodes the geometry of a three-dimensional (3D) ligand conformation into a single 3D point in the space; the second step builds an octree by assigning an octant identifier to every single point in the space under consideration; and the third step performs an octree-based clustering on the reduced conformation space and identifies the most dense octant. We adapt our method for MapReduce and implement it in Hadoop. Load-balancing, fault-tolerance, and scalability in MapReduce allows screening of very large conformation spaces not approachable with traditional clustering methods. We analyze results for docking and crossdocking for a series of HIV protease inhibitors. Our method demonstrates significant improvement over "energy-only" scoring for the accurate identification of native ligand geometries. The advantag...

2012 IEEE 14th International Conference on High Performance Computing and Communication & 2012 IEEE 9th International Conference on Embedded Software and Systems, 2012

Abstract We propose a linear clustering approach for large datasets of molecular geometries produ... more Abstract We propose a linear clustering approach for large datasets of molecular geometries produced by high-throughput molecular dynamics simulations (eg, protein folding and protein-ligand docking simulations). To this scope, we transform each three-dimensional (3D) molecular conformation into a single point in the 3D space reducing the space complexity while still encoding the molecular similarities and geometries. We assign an identifier to each single 3D point mapping a docked ligand, generate a tree from the whole ...

Structure, 2004

with maximal amyloid formation around pH 4.4, where the monomer is the predominant species (Lai e... more with maximal amyloid formation around pH 4.4, where the monomer is the predominant species (Lai et al., Seattle, Washington 98195 1996). The ␤-content (by far-UV CD) of this monomeric form appears to be high, with a signal approximately 75% of that of an engineered, monomeric construct (Lai Summary et al., 1996; Jiang et al., 2001). The tertiary structural changes were more dramatic, as assessed by near-UV The homotetramer of transthyretin (TTR) dissociates CD and fluorescence (Lai et al., 1996; Jiang et al., 2001). into a monomeric amyloidogenic intermediate that This monomeric species has been called an amyloidoself-assembles into amyloid fibrils at low pH. We have genic intermediate. Earlier, this intermediate was reperformed molecular dynamics simulations of monoported not to bind 1-anilino-8-naphthalenesulfonic acid meric TTR at neutral and low pH at physiological (310 (ANS) (Lai et al., 1996), but it has been shown recently K) and very elevated temperature (498 K). In the lowthat it does bind ANS (Jiang et al., 2001). At pH 2, ANS pH simulations at both temperatures, one of the two binding increases and the tertiary structure becomes ␤-sheets (strands CBEF) becomes disrupted, and further disrupted. This lower pH state is incapable of ␣-sheet structure forms in the other sheet (strands forming amyloid. DAGH). ␣-sheet is formed by alternating ␣ L and ␣ R There is a plateau in the denaturation curve of TTR, residues, and it was first proposed by Pauling and as measured by the intrinsic Trp fluorescence over the Corey. Overall, the simulations are in agreement with amyloid-forming pH range (pH 4.3-5.2), suggesting the the available experimental observations, including buildup of an intermediate with tertiary structure distinct solid-state NMR results for a TTR-peptide amyloid. In from that of the native protein (Lai et al., 1996). This addition, they provide a unique explanation for the change in intrinsic fluorescence is due to dynamic results of hydrogen exchange experiments of the amyquenching of Trp41 (Lai et al., 1996). This effect is loidogenic intermediate-results that are difficult to thought to be the result of a structural rearrangement explain with ␤-structure. We propose that ␣-sheet may of the C-strand-loop-D-strand region (hereafter referred represent a key pathological conformation during amto as the CD-loop, residues 40-56) (Figures 2A and 2B) yloidogenesis. (Lai et al., 1996). It has been hypothesized that these two strands become unstructured or detached, revealing the Simulations at 310 K Neutral pH 0-25 5.5

Proteins: Structure, Function, and Bioinformatics, 2010

The arenavirus genome encodes for a Z-protein, which contains a RING domain that coordinates two ... more The arenavirus genome encodes for a Z-protein, which contains a RING domain that coordinates two zinc ions, and has been identified as having several functional roles at various stages of the virus life cycle. Z-protein binds to multiple host proteins and has been directly implicated in the promotion of viral budding, repression of mRNA translation and apoptosis of infected cells. Using homology models of the Z-protein from Lassa strain arenavirus, replica exchange molecular dynamics were employed to refine the structures, which were then subsequently clustered. Population weighted ensembles of low energy cluster representatives were predicted based upon optimal agreement of the chemical shifts computed with the SPARTA program with the experimental NMR chemical shifts. A member of the refined ensemble was indentified to be a potential binder of budding factor Tsg101 based on its correspondence to the structure of the HIV-1 Gag late domain when bound to Tsg101. Members of these ensembles were docked against the crystal structure of human eIF4E translation initiation factor. Two plausible binding modes emerged based upon their agreement with experimental observation, favorable interaction energies and stability during molecular dynamics trajectories. Mutations to Z are proposed that would either inhibit both binding mechanisms or selectively inhibit only one mode. The C-terminal domain conformation of the most populated member of the representative ensemble shielded protein binding recognition motifs for Tsg101 and eIF4E, and represents the most populated state free in solution. We propose that C-terminal flexibility is key for mediating the different functional states of the Z-protein.

Protein Science, 2003

The conformational equilibrium between 3 10-and ␣-helical structure has been studied via high-res... more The conformational equilibrium between 3 10-and ␣-helical structure has been studied via high-resolution NMR spectroscopy by Millhauser and coworkers using the MW peptide Ac-AMAAKAWAAKA AAARA-NH2. Their 750-MHz nuclear Overhauser effect spectroscopy (NOESY) spectra were interpreted to reflect appreciable populations of 3 10-helix throughout the peptide, with the greatest contribution at the N and C termini. The presence of simultaneous ␣N(i,i + 2) and ␣N(i,i + 4) NOE cross-peaks was proposed to represent conformational averaging between 3 10-and ␣-helical structures. In this study, we describe 25-nsec molecular dynamics simulations of the MW peptide at 298 K, using both an 8 Å and a 10 Å force-shifted nonbonded cutoff. The ensemble averages of both simulations are in reasonable agreement with the experimental helical content from circular dichroism (CD), the 3 J HN␣ coupling constants, and the 57 observed NOEs. Analysis of the structures from both simulations revealed very little formation of contiguous i → i + 3 hydrogen bonds (3 10-helix); however, there was a large population of bifurcated i → i + 3 and i → i + 4 ␣-helical hydrogen bonds. In addition, both simulations contained considerable populations of-helix (i → i + 5 hydrogen bonds). Individual turns formed over residues 1-9, which we predict contribute to the intensities of the experimentally observed ␣N(i,i + 2) NOEs. Here we show how sampling of both folded and unfolded structures can provide a structural framework for deconvolution of the conformational contributions to experimental ensemble averages.

Protein Engineering Design and Selection, 2008

Transthyretin (TTR)-containing amyloid fibrils are deposited in cardiac tissue as a natural conse... more Transthyretin (TTR)-containing amyloid fibrils are deposited in cardiac tissue as a natural consequence of aging. A large number of inherited mutations lead to amyloid diseases by accelerating TTR deposition in other organs. Amyloid formation is preceded by a disruption of the quaternary structure of TTR and conformational changes in the monomer. To study conformational changes preceding the formation of amyloid, we performed molecular dynamics simulations of the wild-type monomer, amyloidogenic variants (V30M, L55P, V122I) and a protective variant (T119M) at neutral and low pH. At low pH, the D strand dissociated from the b-sheet to expose the A strand, consistent with experimental studies. In amyloidogenic variants and in the wild-type at low pH, there was a conformational change in the b-sheets into a-sheet via peptide bond flips that was not observed at neutral pH in the wild-type monomer. The same residues participated in conversion in each amyloidogenic variant simulation, originating in the G strand between residues 106 and 109, with accelerated conversion at low pH. The T119M protective variant changed the local conformation of the H strand and suppressed the conversion observed in amyloidogenic variants.

Proceedings of the National Academy of Sciences, 2004

Transthyretin, β 2 -microglobulin, lysozyme, and the prion protein are four of the best-character... more Transthyretin, β 2 -microglobulin, lysozyme, and the prion protein are four of the best-characterized proteins implicated in amyloid disease. Upon partial acid denaturation, these proteins undergo conformational change into an amyloidogenic intermediate that can self-assemble into amyloid fibrils. Many experiments have shown that pH-mediated changes in structure are required for the formation of the amyloidogeneic intermediate, but it has proved impossible to characterize these conformational changes at high resolution using experimental means. To probe these conformational changes at atomic resolution, we have performed molecular dynamics simulations of these proteins at neutral and low pH. In low-pH simulations of all four proteins, we observe the formation of α-pleated sheet secondary structure, which was first proposed by L. Pauling and R. B. Corey [(1951) Proc. Natl. Acad. Sci. USA 37, 251–256]. In all β-sheet proteins, transthyretin and β 2 -microglobulin, α-pleated sheet stru...

Journal of Computational Chemistry, 2013

Molecular docking of small-molecules is an important procedure for computer-aided drug design. Mo... more Molecular docking of small-molecules is an important procedure for computer-aided drug design. Modeling receptor side chain flexibility is often important or even crucial, as it allows the receptor to adopt new conformations as induced by ligand binding. However, the accurate and efficient incorporation of receptor side chain flexibility has proven to be a challenge due to the huge computational complexity required to adequately address this problem. Here we describe a new docking approach with a very fast, graph-based optimization algorithm for assignment of the near-optimal set of residue rotamers. We extensively validate our approach using the 40 DUD target benchmarks commonly used to assess virtual screening performance and demonstrate a large improvement using the developed side chain optimization over rigid receptor docking (average ROC AUC of 0.693 vs. 0.623). Compared to numerous benchmarks, the overall performance is better than nearly all other commonly used procedures. Furthermore, we provide a detailed analysis of the level of receptor flexibility observed in docking results for different classes of residues and elucidate potential avenues for further improvement. V

Journal of Chemical Theory and Computation, 2009

Incorporating receptor flexibility into molecular docking should improve results for flexible pro... more Incorporating receptor flexibility into molecular docking should improve results for flexible proteins. However, the incorporation of explicit all-atom flexibility with molecular dynamics for the entire protein chain may also introduce significant error and "noise" that could decrease docking accuracy and deteriorate the ability of a scoring function to rank native-like poses. We address this apparent paradox by comparing the success of several flexible receptor models in cross-docking and multiple receptor ensemble docking for p38α mitogen-activated protein (MAP) kinase. Explicit all-atom receptor flexibility has been incorporated into a CHARMM-based molecular docking method (CDOCKER) using both molecular dynamics (MD) and torsion angle molecular dynamics (TAMD) for the refinement of predicted protein-ligand binding geometries. These flexible receptor models have been evaluated, and the accuracy and efficiency of TAMD sampling is directly compared to MD sampling. Several flexible receptor models are compared, encompassing flexible side chains, flexible loops, multiple flexible backbone segments, and treatment of the entire chain as flexible. We find that although including side chain and some backbone flexibility is required for improved docking accuracy as expected, docking accuracy also diminishes as additional and unnecessary receptor flexibility is included into the conformational search space. Ensemble docking results demonstrate that including protein flexibility leads to to improved agreement with binding data for 227 active compounds. This comparison also demonstrates that a flexible receptor model enriches high affinity compound identification without significantly increasing the number of false positives from low affinity compounds.

IEEE Engineering in Medicine and Biology Magazine, 2009

In biological systems, the binding of small molecule ligands to proteins is a crucial process for... more In biological systems, the binding of small molecule ligands to proteins is a crucial process for almost every aspect of biochemistry and molecular biology. Enzymes are proteins that function by catalyzing specific biochemical reactions that convert reactants into products. Complex organisms are typically composed of cells in which thousands of enzymes participate in complex and interconnected biochemical pathways. Some enzymes serve as sequential steps in specific pathways (such as energy metabolism), while others function to regulate entire pathways and cellular functions [1]. Small molecule ligands can be designed to bind to a specific enzyme and inhibit the biochemical reaction. Inhibiting the activity of key enzymes may result in the entire biochemical pathways being turned on or off [2], [3]. Many small molecule drugs marketed today function in this generic way as enzyme inhibitors. If research identifies a specific enzyme as being crucial to the progress of disease, then this enzyme may be targeted with an inhibitor, which may slow down or reverse the progress of disease. In this way, enzymes are targeted from specific pathogens (e.g., virus, bacteria, fungi) for infectious diseases [4], [5], and human enzymes are targeted for noninfectious diseases such as cardiovascular disease, cancer, diabetes, and neurodegenerative diseases [6].

Biochemistry, 2005

The correct treatment of van der Waals and electrostatic nonbonded interactions in molecular forc... more The correct treatment of van der Waals and electrostatic nonbonded interactions in molecular force fields is essential for performing realistic molecular dynamics (MD) simulations of solvated polypeptides. The most computationally tractable treatment of nonbonded interactions in MD utilizes a spherical distance cutoff (typically, 8-12 Å) to reduce the number of pairwise interactions. In this work, we assess three spherical atom-based cutoff approaches for use with all-atom explicit solvent MD: abrupt truncation, a CHARMM-style electrostatic shift truncation, and our own force-shifted truncation. The chosen system for this study is an end-capped 17-residue alanine-based R-helical peptide, selected because of its use in previous computational and experimental studies. We compare the time-averaged helical content calculated from these MD trajectories with experiment. We also examine the effect of varying the cutoff treatment and distance on energy conservation. We find that the abrupt truncation approach is pathological in its inability to conserve energy. The CHARMM-style shift truncation performs quite well but suffers from energetic instability. On the other hand, the force-shifted spherical cutoff method conserves energy, correctly predicts the experimental helical content, and shows convergence in simulation statistics as the cutoff is increased. This work demonstrates that by using proper and rigorous techniques, it is possible to correctly model polypeptide dynamics in solution with a spherical cutoff. The inherent computational advantage of spherical cutoffs over Ewald summation (and related) techniques is essential in accessing longer MD time scales.

Proceedings of the First ACM International Conference on Bioinformatics and Computational Biology, 2010

Docking simulations are commonly used to understand drug binding and require the search of a larg... more Docking simulations are commonly used to understand drug binding and require the search of a large space of proteinligand conformations. Cloud and volunteer computing enable computationally expensive docking simulations at a rate never seen before but at the same time require scientists to deal with larger datasets. When analysing these datasets, a common practice is to reduce the resulting number of candidates up to 10 to 100 conformations based on energy values and then leave the scientists with the tedious task of subjectively selecting a possible near-native ligand. Scientists normally perform this task manually by using visual tools. Not only the manual process still depends on inaccurate energy scoring but also can be highly error-prone. The contributions of this paper are twofold: First, we address the problem of extensively searching large spaces of protein-ligand docking conformations, supported by the volunteer computing project Docking@Home (D@H). Second, we address the problem of accurately, and automatically, selecting near-native ligand conformations from the large number of D@H results by using a probabilistic hierarchical clustering based on ligand geometry. Our method holds up even when we test for a search that is not biased by starting from near-native ligand conformations and clearly outperforms energy-based scoring methods.

The Biochemical journal, Jan 22, 2015

The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signal... more The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signaling and regulate cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (T308 in Akt1) in an ATP-dependent manner. In this report, we describe a distinct mechanism to control T308 dephosphorylation, and thus Akt deactivation, that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal MTORC2 phosphorylation site (Ser473) increased phosphatase resistance of the phosphorylated activation loop (pT308) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent T308 phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydropho...

Circulation, 2014

Introduction: The Akt kinase isoforms (Akt1, Akt2 and Akt3) are activated downstream of the insul... more Introduction: The Akt kinase isoforms (Akt1, Akt2 and Akt3) are activated downstream of the insulin receptor, and exert unique effects on cell growth, survival and metabolism. We recently described a novel ATP-dependent "dephosphorylation-resistance cage" in Akt kinases that controls access of cellular phosphatases to dephosphorylate the Akt activation loop (T308 in Akt1). Hypothesis: Here, we describe that, when Akt is localized at the cell membrane, intramolecular interactions of Akt C-terminal sequences with the hydrophobic groove of the kinase domain protect the phosphorylated activation loop (pT308 in Akt1) from cellular phosphatases. Methods and Results: Charged amino acid replacements of Akt C-terminal phosphorylation sites regulated by MTORC2 (Ser473) and by cyclin-dependent kinase 2 (Ser477) increased phospho-T308 protection from cellular phosphatases regardless of subcellular location. Functionally, these phosphatase-resistant Akt variants were refractory to cera...

Journal of Computational Chemistry, 2017

Following insights from recent crystal structures of the muscarinic acetylcholine receptor, bindi... more Following insights from recent crystal structures of the muscarinic acetylcholine receptor, binding modes of Positive Allosteric Modulators (PAMs) were predicted under the assumption that PAMs should bind to the extracellular surface of the active state. A series of well-characterized PAMs for adenosine (A 1 R, A 2A R, A 3 R) and muscarinic acetylcholine (M 1 R, M 5 R) receptors were modeled using both rigid and flexible receptor CHARMM-based molecular docking. Studies of adenosine receptors investigated the molecular basis of the probe-dependence of PAM activity by modeling in complex with specific agonist radioligands. Consensus binding modes map common pharmacophore features of several chemical series to specific binding interactions. These models provide a rationalization of how PAM binding slows agonist radioligand dissociation kinetics. M 1 R PAMs were predicted to bind in the analogous M 2 R PAM LY2119620 binding site. The M 5 R NAM (ML-375) was predicted to bind in the PAM (ML-380) binding site with a unique inducedfit receptor conformation.

The Biochemical journal, Jan 22, 2015

The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signal... more The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signaling and regulate cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (T308 in Akt1) in an ATP-dependent manner. In this report, we describe a distinct mechanism to control T308 dephosphorylation, and thus Akt deactivation, that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal MTORC2 phosphorylation site (Ser473) increased phosphatase resistance of the phosphorylated activation loop (pT308) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent T308 phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydropho...

We present a scalable and accurate method for classifying protein-ligand binding geometries in mo... more We present a scalable and accurate method for classifying protein-ligand binding geometries in molecular docking. Our method is a three-step process: the first step encodes the geometry of a three-dimensional (3D) ligand conformation into a single 3D point in the space; the second step builds an octree by assigning an octant identifier to every single point in the space under consideration; and the third step performs an octree-based clustering on the reduced conformation space and identifies the most dense octant. We adapt our method for MapReduce and implement it in Hadoop. Load-balancing, fault-tolerance, and scalability in MapReduce allows screening of very large conformation spaces not approachable with traditional clustering methods. We analyze results for docking and crossdocking for a series of HIV protease inhibitors. Our method demonstrates significant improvement over "energy-only" scoring for the accurate identification of native ligand geometries. The advantag...

2012 IEEE 14th International Conference on High Performance Computing and Communication & 2012 IEEE 9th International Conference on Embedded Software and Systems, 2012

Abstract We propose a linear clustering approach for large datasets of molecular geometries produ... more Abstract We propose a linear clustering approach for large datasets of molecular geometries produced by high-throughput molecular dynamics simulations (eg, protein folding and protein-ligand docking simulations). To this scope, we transform each three-dimensional (3D) molecular conformation into a single point in the 3D space reducing the space complexity while still encoding the molecular similarities and geometries. We assign an identifier to each single 3D point mapping a docked ligand, generate a tree from the whole ...

Structure, 2004

with maximal amyloid formation around pH 4.4, where the monomer is the predominant species (Lai e... more with maximal amyloid formation around pH 4.4, where the monomer is the predominant species (Lai et al., Seattle, Washington 98195 1996). The ␤-content (by far-UV CD) of this monomeric form appears to be high, with a signal approximately 75% of that of an engineered, monomeric construct (Lai Summary et al., 1996; Jiang et al., 2001). The tertiary structural changes were more dramatic, as assessed by near-UV The homotetramer of transthyretin (TTR) dissociates CD and fluorescence (Lai et al., 1996; Jiang et al., 2001). into a monomeric amyloidogenic intermediate that This monomeric species has been called an amyloidoself-assembles into amyloid fibrils at low pH. We have genic intermediate. Earlier, this intermediate was reperformed molecular dynamics simulations of monoported not to bind 1-anilino-8-naphthalenesulfonic acid meric TTR at neutral and low pH at physiological (310 (ANS) (Lai et al., 1996), but it has been shown recently K) and very elevated temperature (498 K). In the lowthat it does bind ANS (Jiang et al., 2001). At pH 2, ANS pH simulations at both temperatures, one of the two binding increases and the tertiary structure becomes ␤-sheets (strands CBEF) becomes disrupted, and further disrupted. This lower pH state is incapable of ␣-sheet structure forms in the other sheet (strands forming amyloid. DAGH). ␣-sheet is formed by alternating ␣ L and ␣ R There is a plateau in the denaturation curve of TTR, residues, and it was first proposed by Pauling and as measured by the intrinsic Trp fluorescence over the Corey. Overall, the simulations are in agreement with amyloid-forming pH range (pH 4.3-5.2), suggesting the the available experimental observations, including buildup of an intermediate with tertiary structure distinct solid-state NMR results for a TTR-peptide amyloid. In from that of the native protein (Lai et al., 1996). This addition, they provide a unique explanation for the change in intrinsic fluorescence is due to dynamic results of hydrogen exchange experiments of the amyquenching of Trp41 (Lai et al., 1996). This effect is loidogenic intermediate-results that are difficult to thought to be the result of a structural rearrangement explain with ␤-structure. We propose that ␣-sheet may of the C-strand-loop-D-strand region (hereafter referred represent a key pathological conformation during amto as the CD-loop, residues 40-56) (Figures 2A and 2B) yloidogenesis. (Lai et al., 1996). It has been hypothesized that these two strands become unstructured or detached, revealing the Simulations at 310 K Neutral pH 0-25 5.5

Proteins: Structure, Function, and Bioinformatics, 2010

The arenavirus genome encodes for a Z-protein, which contains a RING domain that coordinates two ... more The arenavirus genome encodes for a Z-protein, which contains a RING domain that coordinates two zinc ions, and has been identified as having several functional roles at various stages of the virus life cycle. Z-protein binds to multiple host proteins and has been directly implicated in the promotion of viral budding, repression of mRNA translation and apoptosis of infected cells. Using homology models of the Z-protein from Lassa strain arenavirus, replica exchange molecular dynamics were employed to refine the structures, which were then subsequently clustered. Population weighted ensembles of low energy cluster representatives were predicted based upon optimal agreement of the chemical shifts computed with the SPARTA program with the experimental NMR chemical shifts. A member of the refined ensemble was indentified to be a potential binder of budding factor Tsg101 based on its correspondence to the structure of the HIV-1 Gag late domain when bound to Tsg101. Members of these ensembles were docked against the crystal structure of human eIF4E translation initiation factor. Two plausible binding modes emerged based upon their agreement with experimental observation, favorable interaction energies and stability during molecular dynamics trajectories. Mutations to Z are proposed that would either inhibit both binding mechanisms or selectively inhibit only one mode. The C-terminal domain conformation of the most populated member of the representative ensemble shielded protein binding recognition motifs for Tsg101 and eIF4E, and represents the most populated state free in solution. We propose that C-terminal flexibility is key for mediating the different functional states of the Z-protein.

Protein Science, 2003

The conformational equilibrium between 3 10-and ␣-helical structure has been studied via high-res... more The conformational equilibrium between 3 10-and ␣-helical structure has been studied via high-resolution NMR spectroscopy by Millhauser and coworkers using the MW peptide Ac-AMAAKAWAAKA AAARA-NH2. Their 750-MHz nuclear Overhauser effect spectroscopy (NOESY) spectra were interpreted to reflect appreciable populations of 3 10-helix throughout the peptide, with the greatest contribution at the N and C termini. The presence of simultaneous ␣N(i,i + 2) and ␣N(i,i + 4) NOE cross-peaks was proposed to represent conformational averaging between 3 10-and ␣-helical structures. In this study, we describe 25-nsec molecular dynamics simulations of the MW peptide at 298 K, using both an 8 Å and a 10 Å force-shifted nonbonded cutoff. The ensemble averages of both simulations are in reasonable agreement with the experimental helical content from circular dichroism (CD), the 3 J HN␣ coupling constants, and the 57 observed NOEs. Analysis of the structures from both simulations revealed very little formation of contiguous i → i + 3 hydrogen bonds (3 10-helix); however, there was a large population of bifurcated i → i + 3 and i → i + 4 ␣-helical hydrogen bonds. In addition, both simulations contained considerable populations of-helix (i → i + 5 hydrogen bonds). Individual turns formed over residues 1-9, which we predict contribute to the intensities of the experimentally observed ␣N(i,i + 2) NOEs. Here we show how sampling of both folded and unfolded structures can provide a structural framework for deconvolution of the conformational contributions to experimental ensemble averages.

Protein Engineering Design and Selection, 2008

Transthyretin (TTR)-containing amyloid fibrils are deposited in cardiac tissue as a natural conse... more Transthyretin (TTR)-containing amyloid fibrils are deposited in cardiac tissue as a natural consequence of aging. A large number of inherited mutations lead to amyloid diseases by accelerating TTR deposition in other organs. Amyloid formation is preceded by a disruption of the quaternary structure of TTR and conformational changes in the monomer. To study conformational changes preceding the formation of amyloid, we performed molecular dynamics simulations of the wild-type monomer, amyloidogenic variants (V30M, L55P, V122I) and a protective variant (T119M) at neutral and low pH. At low pH, the D strand dissociated from the b-sheet to expose the A strand, consistent with experimental studies. In amyloidogenic variants and in the wild-type at low pH, there was a conformational change in the b-sheets into a-sheet via peptide bond flips that was not observed at neutral pH in the wild-type monomer. The same residues participated in conversion in each amyloidogenic variant simulation, originating in the G strand between residues 106 and 109, with accelerated conversion at low pH. The T119M protective variant changed the local conformation of the H strand and suppressed the conversion observed in amyloidogenic variants.

Proceedings of the National Academy of Sciences, 2004

Transthyretin, β 2 -microglobulin, lysozyme, and the prion protein are four of the best-character... more Transthyretin, β 2 -microglobulin, lysozyme, and the prion protein are four of the best-characterized proteins implicated in amyloid disease. Upon partial acid denaturation, these proteins undergo conformational change into an amyloidogenic intermediate that can self-assemble into amyloid fibrils. Many experiments have shown that pH-mediated changes in structure are required for the formation of the amyloidogeneic intermediate, but it has proved impossible to characterize these conformational changes at high resolution using experimental means. To probe these conformational changes at atomic resolution, we have performed molecular dynamics simulations of these proteins at neutral and low pH. In low-pH simulations of all four proteins, we observe the formation of α-pleated sheet secondary structure, which was first proposed by L. Pauling and R. B. Corey [(1951) Proc. Natl. Acad. Sci. USA 37, 251–256]. In all β-sheet proteins, transthyretin and β 2 -microglobulin, α-pleated sheet stru...

Journal of Computational Chemistry, 2013

Molecular docking of small-molecules is an important procedure for computer-aided drug design. Mo... more Molecular docking of small-molecules is an important procedure for computer-aided drug design. Modeling receptor side chain flexibility is often important or even crucial, as it allows the receptor to adopt new conformations as induced by ligand binding. However, the accurate and efficient incorporation of receptor side chain flexibility has proven to be a challenge due to the huge computational complexity required to adequately address this problem. Here we describe a new docking approach with a very fast, graph-based optimization algorithm for assignment of the near-optimal set of residue rotamers. We extensively validate our approach using the 40 DUD target benchmarks commonly used to assess virtual screening performance and demonstrate a large improvement using the developed side chain optimization over rigid receptor docking (average ROC AUC of 0.693 vs. 0.623). Compared to numerous benchmarks, the overall performance is better than nearly all other commonly used procedures. Furthermore, we provide a detailed analysis of the level of receptor flexibility observed in docking results for different classes of residues and elucidate potential avenues for further improvement. V

Journal of Chemical Theory and Computation, 2009

Incorporating receptor flexibility into molecular docking should improve results for flexible pro... more Incorporating receptor flexibility into molecular docking should improve results for flexible proteins. However, the incorporation of explicit all-atom flexibility with molecular dynamics for the entire protein chain may also introduce significant error and "noise" that could decrease docking accuracy and deteriorate the ability of a scoring function to rank native-like poses. We address this apparent paradox by comparing the success of several flexible receptor models in cross-docking and multiple receptor ensemble docking for p38α mitogen-activated protein (MAP) kinase. Explicit all-atom receptor flexibility has been incorporated into a CHARMM-based molecular docking method (CDOCKER) using both molecular dynamics (MD) and torsion angle molecular dynamics (TAMD) for the refinement of predicted protein-ligand binding geometries. These flexible receptor models have been evaluated, and the accuracy and efficiency of TAMD sampling is directly compared to MD sampling. Several flexible receptor models are compared, encompassing flexible side chains, flexible loops, multiple flexible backbone segments, and treatment of the entire chain as flexible. We find that although including side chain and some backbone flexibility is required for improved docking accuracy as expected, docking accuracy also diminishes as additional and unnecessary receptor flexibility is included into the conformational search space. Ensemble docking results demonstrate that including protein flexibility leads to to improved agreement with binding data for 227 active compounds. This comparison also demonstrates that a flexible receptor model enriches high affinity compound identification without significantly increasing the number of false positives from low affinity compounds.

IEEE Engineering in Medicine and Biology Magazine, 2009

In biological systems, the binding of small molecule ligands to proteins is a crucial process for... more In biological systems, the binding of small molecule ligands to proteins is a crucial process for almost every aspect of biochemistry and molecular biology. Enzymes are proteins that function by catalyzing specific biochemical reactions that convert reactants into products. Complex organisms are typically composed of cells in which thousands of enzymes participate in complex and interconnected biochemical pathways. Some enzymes serve as sequential steps in specific pathways (such as energy metabolism), while others function to regulate entire pathways and cellular functions [1]. Small molecule ligands can be designed to bind to a specific enzyme and inhibit the biochemical reaction. Inhibiting the activity of key enzymes may result in the entire biochemical pathways being turned on or off [2], [3]. Many small molecule drugs marketed today function in this generic way as enzyme inhibitors. If research identifies a specific enzyme as being crucial to the progress of disease, then this enzyme may be targeted with an inhibitor, which may slow down or reverse the progress of disease. In this way, enzymes are targeted from specific pathogens (e.g., virus, bacteria, fungi) for infectious diseases [4], [5], and human enzymes are targeted for noninfectious diseases such as cardiovascular disease, cancer, diabetes, and neurodegenerative diseases [6].

Biochemistry, 2005

The correct treatment of van der Waals and electrostatic nonbonded interactions in molecular forc... more The correct treatment of van der Waals and electrostatic nonbonded interactions in molecular force fields is essential for performing realistic molecular dynamics (MD) simulations of solvated polypeptides. The most computationally tractable treatment of nonbonded interactions in MD utilizes a spherical distance cutoff (typically, 8-12 Å) to reduce the number of pairwise interactions. In this work, we assess three spherical atom-based cutoff approaches for use with all-atom explicit solvent MD: abrupt truncation, a CHARMM-style electrostatic shift truncation, and our own force-shifted truncation. The chosen system for this study is an end-capped 17-residue alanine-based R-helical peptide, selected because of its use in previous computational and experimental studies. We compare the time-averaged helical content calculated from these MD trajectories with experiment. We also examine the effect of varying the cutoff treatment and distance on energy conservation. We find that the abrupt truncation approach is pathological in its inability to conserve energy. The CHARMM-style shift truncation performs quite well but suffers from energetic instability. On the other hand, the force-shifted spherical cutoff method conserves energy, correctly predicts the experimental helical content, and shows convergence in simulation statistics as the cutoff is increased. This work demonstrates that by using proper and rigorous techniques, it is possible to correctly model polypeptide dynamics in solution with a spherical cutoff. The inherent computational advantage of spherical cutoffs over Ewald summation (and related) techniques is essential in accessing longer MD time scales.

Proceedings of the First ACM International Conference on Bioinformatics and Computational Biology, 2010

Docking simulations are commonly used to understand drug binding and require the search of a larg... more Docking simulations are commonly used to understand drug binding and require the search of a large space of proteinligand conformations. Cloud and volunteer computing enable computationally expensive docking simulations at a rate never seen before but at the same time require scientists to deal with larger datasets. When analysing these datasets, a common practice is to reduce the resulting number of candidates up to 10 to 100 conformations based on energy values and then leave the scientists with the tedious task of subjectively selecting a possible near-native ligand. Scientists normally perform this task manually by using visual tools. Not only the manual process still depends on inaccurate energy scoring but also can be highly error-prone. The contributions of this paper are twofold: First, we address the problem of extensively searching large spaces of protein-ligand docking conformations, supported by the volunteer computing project Docking@Home (D@H). Second, we address the problem of accurately, and automatically, selecting near-native ligand conformations from the large number of D@H results by using a probabilistic hierarchical clustering based on ligand geometry. Our method holds up even when we test for a search that is not biased by starting from near-native ligand conformations and clearly outperforms energy-based scoring methods.

The Biochemical journal, Jan 22, 2015

The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signal... more The Akt protein kinase, also known as protein kinase B, play key roles in insulin receptor signaling and regulate cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (T308 in Akt1) in an ATP-dependent manner. In this report, we describe a distinct mechanism to control T308 dephosphorylation, and thus Akt deactivation, that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal MTORC2 phosphorylation site (Ser473) increased phosphatase resistance of the phosphorylated activation loop (pT308) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent T308 phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydropho...