Bulletin of the American Physical Society
APS March Meeting 2013
Volume 58, Number 1
Monday–Friday, March 18–22, 2013; Baltimore, Maryland
Session G45: Focus Session: Physics of Protein Aggregation |
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Sponsoring Units: DBIO DPOLY DCP Chair: Daniel Cox, UC Davis Room: Hilton Baltimore Holiday Ballroom 4 |
Tuesday, March 19, 2013 11:15AM - 11:51AM |
G45.00001: Single Molecule Visualization of Protein-DNA Complexes: Watching Machines at Work Invited Speaker: Stephen Kowalczykowski We can now watch individual proteins acting on single molecules of DNA. Such imaging provides unprecedented interrogation of fundamental biophysical processes. Visualization is achieved through the application of two complementary procedures. In one, single DNA molecules are attached to a polystyrene bead and are then captured by an optical trap. The DNA, a worm-like coil, is extended either by the force of solution flow in a micro-fabricated channel, or by capturing the opposite DNA end in a second optical trap. In the second procedure, DNA is attached by one end to a glass surface. The coiled DNA is elongated either by continuous solution flow or by subsequently tethering the opposite end to the surface. Protein action is visualized by fluorescent reporters: fluorescent dyes that bind double-stranded DNA (dsDNA), fluorescent biosensors for single-stranded DNA (ssDNA), or fluorescently-tagged proteins. Individual molecules are imaged using either epifluorescence microscopy or total internal reflection fluorescence (TIRF) microscopy. Using these approaches, we imaged the search for DNA sequence homology conducted by the RecA-ssDNA filament. The manner by which RecA protein finds a single homologous sequence in the genome had remained undefined for almost 30 years. Single-molecule imaging revealed that the search occurs through a mechanism termed ``intersegmental contact sampling,'' in which the randomly coiled structure of DNA is essential for reiterative sampling of DNA sequence identity: an example of parallel processing. In addition, the assembly of RecA filaments on single molecules of single-stranded DNA was visualized. Filament assembly requires nucleation of a protein dimer on DNA, and subsequent growth occurs via monomer addition. Furthermore, we discovered a class of proteins that catalyzed both nucleation and growth of filaments, revealing how the cell controls assembly of this protein-DNA complex. [Preview Abstract] |
Tuesday, March 19, 2013 11:51AM - 12:03PM |
G45.00002: Hydrogen Bonding Motifs in MutSaphla and their response to binding damaged DNA Lacra Negureanu, Freddie Salsbury Over the past decade, there has been a growing interest in studying the binding of damaged DNA to the MutSalpha protein complex. This protein complex, the Msh2/Msh6 complex in humans, is the initial complex that binds mismatched DNA and other DNA defects that occur during replication. This complex has also been shown to bind at least some types of damaged DNA. As a result of this interest, multiple studies have contrasted the interactions of MutSalpha with its normal mismatched substrate and with the interactions of MutsSalpha to DNA damaged by the chemotherapeutic cisplatin. To complement these studies, we examined the interaction between MutSalpha and DNA damaged by carboplatin via all-atom molecular dynamics simulations. These simulations provide evidence for different hydrogen bonding interactions at the protein/DNA and protein/protein interface. The hydrogen bonding motifs found are broadly similar to those found in binding to the adduct from cis-platin, but have distinct differences. These subtle differences may play a role in the way the different damages are signaled by MutS. [Preview Abstract] |
Tuesday, March 19, 2013 12:03PM - 12:15PM |
G45.00003: Parallel Verlet Neighbor List Algorithm for GPU-Optimized MD Simulations Samuel Cho How biomolecules fold and assemble into well-defined structures that correspond to cellular functions is a fundamental problem in biophysics. Molecular dynamics (MD) simulations provide a molecular-resolution physical description of the folding and assembly processes, but the computational demands of the algorithms restrict the size and the timescales one can simulate. In a recent study, we introduced a parallel neighbor list algorithm that was specifically optimized for MD simulations on GPUs. We now analyze the performance of our MD simulation code that incorporates the algorithm, and we observe that the force calculations and the evaluation of the neighbor list and pair lists constitute a majority of the overall execution time. The overall speedup of the GPU-optimized MD simulations as compared to the CPU-optimized version is N-dependent and $\sim$ 30x for the full 70s ribosome (10,219 beads). The pair and neighbor list evaluations have performance speedups of $\sim$ 25x and $\sim$ 55x, respectively. We then make direct comparisons with the performance of our MD simulation code with that of the SOP model implemented in the simulation code of HOOMD, a leading general particle dynamics simulation package that is specifically optimized for GPUs. [Preview Abstract] |
Tuesday, March 19, 2013 12:15PM - 12:27PM |
G45.00004: Coarse-grained Simulations of Protein-Protein Association: Energy Landscape on a Globe Sichun Yang Understanding how proteins interact and associate into large functional complexes is critical in revealing the molecular basis of virtually every biological process in a living cell. Here, a theoretical simulation pipeline using coarse-grained (CG) models with an efficient sampling method is presented from the studies of protein-protein association. A concept of ``energy globe'' is introduced and implemented via the projection of simulation data onto a three-dimensional globe specifying protein-protein orientations and interacting energies. This energy-globe approach has the key advantage of locating and identifying multiple stable conformations that are physically accessible on the energy landscape. Tests on several well-studied protein-protein complexes show that the crystal-like conformation is favorable on the energy landscape even if the landscape is relatively rugged with metastable conformations. Recent applications to CG simulations of nuclear hormone receptors, whose experimental structure are still lacking, have predicted multiple favorable conformations on their corresponding landscapes, thereby providing insight into the cross-talk mechanisms of functional domains in the hormone signaling. [Preview Abstract] |
Tuesday, March 19, 2013 12:27PM - 12:39PM |
G45.00005: Snyder-Robinson Syndrome: Rescuing the Disease-Causing Effect of G56S mutant by Small Molecule Binding Zhe Zhang, Virginie Martiny, David Lagorce, Emil Alexov, Maria Miteva Snyder-Robinson Syndrome (SRS) is an X-linked mental retardation disorder, which is caused by defects in a particular gene coding for the spermine synthase (SMS) protein. Among the missense mutations known to be disease-causing is the G56S, which is positioned at the interface of the SMS homo-dimer. Previous computational and experimental investigations have shown that G56S mutation destabilizes the homo-dimer and thus greatly reduces the SMS enzymatic activity. In this study, we explore the possibility of mitigating the effect of G56S mutation by binding small molecules to suitable pockets around the mutation site. It is done by combined efforts of molecular dynamics simulations and in silico screening. The binding of selected molecules was calculated to fully compensate the effect of the mutation and rescue the wild type dimer affinity. [Preview Abstract] |
Tuesday, March 19, 2013 12:39PM - 12:51PM |
G45.00006: Aggregation of concentrated monoclonal antibody solutions studied by rheology and neutron scattering Maria Monica Castellanos, Jai Pathak, Ralph Colby Protein solutions are studied using rheology and scattering techniques to investigate aggregation. Here we present a monoclonal antibody (mAb) that aggregates after incubation at 40 $^{\circ}$C (below its unfolding temperature), with a decrease in monomer purity of 6{\%} in 10 days. The mAb solution contains surfactant and behaves as a Newtonian fluid when reconstituted into solution from the lyophilized form (before incubation at 40 $^{\circ}$C). In contrast, mAb solutions incubated at 40 $^{\circ}$C for 1 month exhibit shear yielding in torsional bulk rheometers. Interfacial rheology reveals that interfacial properties are controlled by the surfactant, producing a negligible surface contribution to the bulk yield stress. These results provide evidence that protein aggregates formed in the bulk are responsible for the yield stress. Small-angle neutron scattering (SANS) measurements show an increase in intensity at low wavevectors (q \textless\ 4*10$^{-2}$ nm$^{-1})$ that we attribute to protein aggregation, and is not observed in solutions stored at 4 $^{\circ}$C for 3 days before the measurement. This work suggests a correlation between the aggregated state of the protein (stability) and the yield stress from rheology. [Preview Abstract] |
Tuesday, March 19, 2013 12:51PM - 1:03PM |
G45.00007: Enhancing Nucleation rates using Porous Silica Sathish Akella, Seth Fraden The role of nucleants in promoting protein crystal nucleation is an on-going field of research. Porous silica acts as heterogeneous nucleation centers and enhances nucleation rates. For the protein lysozyme there are multiple polymorphs and we demonstrate that porous silica preferentially increases one of the polymorphs. Preliminary studies are presented in which accurate nucleation rates for the different polymorphs as a function of nucleant concentration are obtained through optical microscopy studies of thousands of crystallization trials in identical water-in-oil emulsion drops produced using microfluidics. [Preview Abstract] |
Tuesday, March 19, 2013 1:03PM - 1:15PM |
G45.00008: Solvent-induced size reduction of self-assembled siRNA/copolymer nanoparticles Wei Qu, Juan Wu, Hai-Quan Mao, Erik Luijten Small interfering RNA (siRNA) therapeutics has a demonstrated potential for treating numerous liver diseases. However, traditional polycation vectors used for siRNA delivery typically produce siRNA-containing particles of large size ($>100$~nm), along with high cytotoxicity and low colloidal stability. Inspired by earlier work on nanoparticles for plasmid DNA delivery [1], we graft hydrophilic and biocompatible polyethylene glycol (PEG) blocks to the polycation vector to overcome these limitations. We find that the PEG-grafted polycations result in slightly larger particle size, even though the hydrophilic PEG blocks are expected to hinder the formation of larger aggregates. To explain this observation, we investigate siRNA/copolymer self-assembly via computer simulations of coarse-grained polymer and siRNA models. Our calculations suggest that hydrogen bonding between PEG and the polycation leads to the increased particle size, and that smaller particles can be obtained by inhibiting hydrogen bonding in such system. Subsequent experiments employing solvents of lower polarity indeed lead to particles with smaller size. \\[4pt] [1] X. Jiang \emph{et al.}, Adv. Mater., doi: 10.1002/adma.201202932 [Preview Abstract] |
Tuesday, March 19, 2013 1:15PM - 1:27PM |
G45.00009: Using Nanoscale Substrate Curvature to Control the Dimerization of Surface-Bound Proteins Hilary Paulin, Martin Kurylowicz, Josh Mogyoros, Maximiliano Giuliani, John Dutcher The influence of surface geometry on adsorbed proteins offers new possibilities for controlling quaternary structure by manipulating protein-protein interactions at a surface, with applications that are relevant to protein aggregation, fibrillation, ligand binding and surface catalysis. To understand the effect of surface curvature on the structure of surface-bound proteins, we have used a combination of polystyrene (PS) nanoparticles (NPs) and ultrathin PS films to fabricate chemically pure, hydrophobic surfaces that have nanoscale curvature and are stable in aqueous buffer. We have used Single Molecule Force Spectroscopy (SMFS) to measure the detachment contour lengths$~(L_{c})$ for~beta-lactoglobulin (b-LG) and~alpha-lactalbumin (a-LA) adsorbed onto neighbouring regions of highly curved and flat PS surfaces, allowing us to compare these values~\textit{in situ}~on the same sample.~In general, we measure peaks in the $L_{c}$ distributions corresponding to monomers and dimers. As the curvature of the underlying surface is increased, the population of dimers decreases such that only monomers are observed for b-LG adsorbed onto 25 nm dia NPs. These results indicate that surface curvature provides a new method of manipulating protein-protein interactions and controlling the quaternary structure of adsorbed proteins. [Preview Abstract] |
Tuesday, March 19, 2013 1:27PM - 1:39PM |
G45.00010: Coarse-grained Molecular Dynamics Simulation of Calmodulin-target Interactions Pengzhi Zhang, Qian Wang, Swarnendu Tripathi, Margaret Cheung Calmodulin (CaM) is a ubiquitous small protein playing an important role in Ca$^{\mathrm{2+}}$ signaling in eukaryotic cells, which can bind and regulate hundreds of target enzymes in the presence of Ca$^{\mathrm{2+}}$. Although the binding process is known to be diffusion controlled, however, due to the flexibility of CaM, methodology that provides molecular insights on target binding and recognition. In this study, Brownian dynamics simulations were used to mimic the process Ca$^{\mathrm{2+}}$-bound CaM binds with two target peptides: CaMKI and CaMKII. Using an experimentally-analogous criterion of number of contacts between targets and a specific residue of CaM to define the encounter complexes and to calculate the association rates, we are able to reveal the molecular reason why CaM-CaMKI has twice the rate of CaM-CaMKII while the numbers of amino acids are similar. [Preview Abstract] |
Tuesday, March 19, 2013 1:39PM - 1:51PM |
G45.00011: Linear and Nonlinear Microrheology of Interfacial Protein Layers Daniel Allan, Daniel Firester, Victor Allard, Daniel Reich, Robert Leheny Proteins can adsorb to the air-water interface to form a robust layer. As protein accretes and a layer forms, we monitor the layer's shear rheology employing both passive and active microrheology. Measurements of the linear rheology, using multiple-particle-tracking techniques, show a transition from a viscous to elastic interface with increasing layer age. Active measurements of the nonlinear rheology, in which ferromagnetic nanowires at the interface rotate in response to magnetic torques, show that the protein layers behave quantitatively like a Hershel-Bulkley fluid. We interpret these observations in terms of mechanisms of layer formation and protein interactions at the interface. [Preview Abstract] |
Tuesday, March 19, 2013 1:51PM - 2:03PM |
G45.00012: Contributions of equilibrium and non-equilibrium clusters to viscosity in concentrated protein solutions Prasad Sarangapani, Steven Hudson, Jai Pathak, Kalman Migler Equilibrium and non-equilibrium clustering are ubiquitous phenomena in soft matter physics and are typically observed in systems ranging from colloidal suspensions to monoclonal antibodies (mAbs). Such phenomena are central to understanding and preventing irreversible aggregation in addition to controlling viscosity challenges related to formulation and drug delivery of protein therapeutics. Curiously, little work has been done in exploring the cluster size dependence of low-shear viscosity and intrinsic viscosity in protein solutions in a controlled manner. In this work, we carefully control cluster size of reversible and irreversible clusters formed by globular proteins or monoclonal antibodies over a concentration range of 2 mg/mL-500 mg/mL and pH from 3-9. We find a marked dependence of low-shear viscosity on cluster size using custom-designed silicon-based microfluidic viscometers. Measurements of cluster sizes using static light scattering reveal a correlation of low shear viscosity as well as intrinsic viscosity with the average cluster size. We model the composition dependence of viscosity for the case of equilibrium and non-equilibrium clusters using an adaptation of a model recently presented by Minton for protein mixtures. [Preview Abstract] |
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