Publications by authors named "Emre Brookes"

19 Publications

  • Page 1 of 1

Mechanism of NanR gene repression and allosteric induction of bacterial sialic acid metabolism.

Nat Commun 2021 03 31;12(1):1988. Epub 2021 Mar 31.

Biomolecular Interaction Centre and School of Biological Sciences, University of Canterbury, Christchurch, New Zealand.

Bacteria respond to environmental changes by inducing transcription of some genes and repressing others. Sialic acids, which coat human cell surfaces, are a nutrient source for pathogenic and commensal bacteria. The Escherichia coli GntR-type transcriptional repressor, NanR, regulates sialic acid metabolism, but the mechanism is unclear. Here, we demonstrate that three NanR dimers bind a (GGTATA)-repeat operator cooperatively and with high affinity. Single-particle cryo-electron microscopy structures reveal the DNA-binding domain is reorganized to engage DNA, while three dimers assemble in close proximity across the (GGTATA)-repeat operator. Such an interaction allows cooperative protein-protein interactions between NanR dimers via their N-terminal extensions. The effector, N-acetylneuraminate, binds NanR and attenuates the NanR-DNA interaction. The crystal structure of NanR in complex with N-acetylneuraminate reveals a domain rearrangement upon N-acetylneuraminate binding to lock NanR in a conformation that weakens DNA binding. Our data provide a molecular basis for the regulation of bacterial sialic acid metabolism.
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http://dx.doi.org/10.1038/s41467-021-22253-6DOI Listing
March 2021

Measuring compressibility in the optima AUC™ analytical ultracentrifuge.

Eur Biophys J 2020 Dec 24;49(8):711-718. Epub 2020 Nov 24.

Department of Chemistry, University of Montana, Missoula, MT, USA.

A method is described to accurately measure the compressibility of liquids using an analytical ultracentrifuge. The method makes use of very large pressure gradients, which can be generated in the analytical ultracentrifuge at high speeds to induce a maximum compression signal. Taking advantage of the new Optima AUC, which offers 10 micron radial resolution, a novel calibration centerpiece for measuring rotor stretch, and a speed-ramping procedure, even the weak compressibility of liquids like water, typically considered to be incompressible, could be detected. A model using the standard expression for the secant-average bulk modulus describing the relative compression of a liquid in the analytical ultracentrifuge is derived. The model is a function of the loading volume and the hydrostatic pressure generated in the analytical ultracentrifuge, as well as the secant-average bulk modulus. The compressibility of water and toluene were measured and the linear secant-average bulk modulus and meniscus positions were fitted. In addition to the measurement of the compressibility of liquids, applications for this method include an improved prediction of boundary conditions for multi-speed analytical ultracentrifugation experiments to better describe highly heterogeneous systems with analytical speed-ramping procedures, and the prediction of radius-dependent density variations.
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http://dx.doi.org/10.1007/s00249-020-01482-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7751350PMC
December 2020

BEES: Bayesian Ensemble Estimation from SAS.

Biophys J 2019 08 18;117(3):399-407. Epub 2019 Jul 18.

Department of Physics and the Center for Molecular Study of Condensed Soft Matter, Illinois Institute of Technology, Chicago, Illinois.

Many biomolecular complexes exist in a flexible ensemble of states in solution that is necessary to perform their biological function. Small-angle scattering (SAS) measurements are a popular method for characterizing these flexible molecules because of their relative ease of use and their ability to simultaneously probe the full ensemble of states. However, SAS data is typically low dimensional and difficult to interpret without the assistance of additional structural models. In theory, experimental SAS curves can be reconstituted from a linear combination of theoretical models, although this procedure carries a significant risk of overfitting the inherently low-dimensional SAS data. Previously, we developed a Bayesian-based method for fitting ensembles of model structures to experimental SAS data that rigorously avoids overfitting. However, we have found that these methods can be difficult to incorporate into typical SAS modeling workflows, especially for users that are not experts in computational modeling. To this end, we present the Bayesian Ensemble Estimation from SAS (BEES) program. Two forks of BEES are available, the primary one existing as a module for the SASSIE web server and a developmental version that is a stand-alone Python program. BEES allows users to exhaustively sample ensemble models constructed from a library of theoretical states and to interactively analyze and compare each model's performance. The fitting routine also allows for secondary data sets to be supplied, thereby simultaneously fitting models to both SAS data as well as orthogonal information. The flexible ensemble of K63-linked ubiquitin trimers is presented as an example of BEES' capabilities.
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http://dx.doi.org/10.1016/j.bpj.2019.06.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6698195PMC
August 2019

High-Resolution Asymmetrical Flow Field-Flow Fractionation Data Evaluation via Richardson-Lucy-Based Fractogram Correction.

Anal Chem 2018 12 13;90(23):13978-13986. Epub 2018 Nov 13.

Coriolis Pharma Research GmbH, Frauenhoferstrasse 18b , 82152 Martinsried , Germany.

Asymmetrical flow field-flow fractionation (AF4) is a chromatographic separation technique that can be used for a broad range of particles or macromolecules. As an orthogonal method to size exclusion chromatography (SEC) with a much broader separation size range (1-800 nm) AF4 is gaining importance. However, the data evaluation capacities are far behind in comparison to other techniques like analytical ultracentrifugation (AUC). A program for evaluation of data from AF4 with a coupled multiangle laser light scattering (MALLS) detector was developed that allows the determination of the distributions of diffusion coefficients ( D), hydrodynamic radii ( R), molecular weights ( M), and relative concentrations (RC) of the obtained species. In addition, two algorithms to remove broadening effects via deconvolution were implemented and tested for their validity. The first is an extension of the known diffusion broadening correction applying the entire diffusion coefficient distribution instead of a single diffusion coefficient. The second applies the Richardson-Lucy algorithm for the deconvolution of overlapping signals from stars in astronomy. This program allows a reproducible strong enhancement of the fractogram resolution allowing for entire baseline separations of proteins. The comparison of the values for M determined by a partial Zimm plot from each data point of the original fractogram and the deconvolved results shows that especially the Richardson-Lucy algorithm maintains a high degree of data robustness.
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http://dx.doi.org/10.1021/acs.analchem.8b03483DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6643961PMC
December 2018

Two-dimensional grid optimization for sedimentation velocity analysis in the analytical ultracentrifuge.

Eur Biophys J 2018 Oct 17;47(7):837-844. Epub 2018 May 17.

Department of Biochemistry and Structural Biology, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX, 78229-3901, USA.

Sedimentation velocity experiments performed in the analytical ultracentrifuge are modeled using finite-element solutions of the Lamm equation. During modeling, three fundamental parameters are optimized: the sedimentation coefficients, the diffusion coefficients, and the partial concentrations of all solutes present in a mixture. A general modeling approach consists of fitting the partial concentrations of solutes defined in a two-dimensional grid of sedimentation and diffusion coefficient combinations that cover the range of possible solutes for a given mixture. An increasing number of grid points increase the resolution of the model produced by the subsequent analysis, with denser grids giving rise to a very large system of equations. Here, we evaluate the efficiency and resolution of several regular grids and show that traditionally defined grids tend to provide inadequate coverage in one region of the grid, while at the same time being computationally wasteful in other sections of the grid. We describe a rapid and systematic approach for generating efficient two-dimensional analysis grids that balance optimal information content and model resolution for a given signal-to-noise ratio with improved calculation efficiency. These findings are general and apply to one- and two-dimensional grids, although they no longer represent regular grids. We provide a recipe for an improved grid-point spacing in both directions which eliminates unnecessary points, while at the same time providing a more uniform resolution that can be scaled based on the stochastic noise in the experimental data.
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http://dx.doi.org/10.1007/s00249-018-1309-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158092PMC
October 2018

Recent advances in the UltraScan SOlution MOdeller (US-SOMO) hydrodynamic and small-angle scattering data analysis and simulation suite.

Eur Biophys J 2018 Oct 28;47(7):855-864. Epub 2018 Mar 28.

Biopolimeri e Proteomica, Ospedale Policlinico San Martino, Largo R. Benzi 10, Genova, 16132, Italy.

The UltraScan SOlution MOdeller (US-SOMO) is a comprehensive, public domain, open-source suite of computer programs centred on hydrodynamic modelling and small-angle scattering (SAS) data analysis and simulation. We describe here the advances that have been implemented since its last official release (#3087, 2017), which are available from release #3141 for Windows, Linux and Mac operating systems. A major effort has been the transition from the legacy Qt3 cross platform software development and user interface library to the modern Qt5 release. Apart from improved graphical support, this has allowed the direct implementation of the newest, almost two-orders of magnitude faster version of the ZENO hydrodynamic computation algorithm for all operating systems. Coupled with the SoMo-generated bead models with overlaps, ZENO provides the most accurate translational friction computations from atomic-level structures available (Rocco and Byron Eur Biophys J 44:417-431, 2015a), with computational times comparable with or faster than those of other methods. In addition, it has allowed us to introduce the direct representation of each atom in a structure as a (hydrated) bead, opening interesting new modelling possibilities. In the small-angle scattering (SAS) part of the suite, an indirect Fourier transform Bayesian algorithm has been implemented for the computation of the pairwise distance distribution function from SAS data. Finally, the SAS HPLC module, recently upgraded with improved baseline correction and Gaussian decomposition of not baseline-resolved peaks and with advanced statistical evaluation tools (Brookes et al. J Appl Cryst 49:1827-1841, 2016), now allows automatic top-peak frame selection and averaging.
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http://dx.doi.org/10.1007/s00249-018-1296-0DOI Listing
October 2018

Atomistic modelling of scattering data in the Collaborative Computational Project for Small Angle Scattering (CCP-SAS).

J Appl Crystallogr 2016 Dec 14;49(Pt 6):1861-1875. Epub 2016 Oct 14.

Center for Neutron Research, National Institute of Standards and Technology , Gaithersburg, MD 20899-8562, USA.

The capabilities of current computer simulations provide a unique opportunity to model small-angle scattering (SAS) data at the atomistic level, and to include other structural constraints ranging from molecular and atomistic energetics to crystallography, electron microscopy and NMR. This extends the capabilities of solution scattering and provides deeper insights into the physics and chemistry of the systems studied. Realizing this potential, however, requires integrating the experimental data with a new generation of modelling software. To achieve this, the CCP-SAS collaboration (http://www.ccpsas.org/) is developing open-source, high-throughput and user-friendly software for the atomistic and coarse-grained molecular modelling of scattering data. Robust state-of-the-art molecular simulation engines and molecular dynamics and Monte Carlo force fields provide constraints to the solution structure inferred from the small-angle scattering data, which incorporates the known physical chemistry of the system. The implementation of this software suite involves a tiered approach in which provides the deployment infrastructure for running applications on both standard and high-performance computing hardware, and provides a workflow framework into which modules can be plugged to prepare structures, carry out simulations, calculate theoretical scattering data and compare results with experimental data. produces the accessible web-based front end termed , and and also make community SAS codes available. Applications are illustrated by case studies: (i) inter-domain flexibility in two- to six-domain proteins as exemplified by HIV-1 Gag, MASP and ubiquitin; (ii) the hinge conformation in human IgG2 and IgA1 antibodies; (iii) the complex formed between a hexameric protein Hfq and mRNA; and (iv) synthetic 'bottlebrush' polymers.
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http://dx.doi.org/10.1107/S160057671601517XDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5139988PMC
December 2016

HPLC-SAXS module: dealing with capillary fouling and extraction of pure component patterns from poorly resolved SEC-SAXS data.

J Appl Crystallogr 2016 Oct 1;49(Pt 5):1827-1841. Epub 2016 Sep 1.

SWING Beamline, Synchrotron SOLEIL, L'Orme des Merisiers, BP48, Saint-Aubin, Gif-sur-Yvette, F-91192, France.

Size-exclusion chromatography coupled with SAXS (small-angle X-ray scattering), often performed using a flow-through capillary, should allow direct collection of monodisperse sample data. However, capillary fouling issues and non-baseline-resolved peaks can hamper its efficacy. The solution modeler () HPLC-SAXS (high-performance liquid chromatography coupled with SAXS) module provides a comprehensive framework to analyze such data, starting with a simple linear baseline correction and symmetrical Gaussian decomposition tools [Brookes, Pérez, Cardinali, Profumo, Vachette & Rocco (2013 ▸). , 1823-1833]. In addition to several new features, substantial improvements to both routines have now been implemented, comprising the evaluation of outcomes by advanced statistical tools. The novel integral baseline-correction procedure is based on the more sound assumption that the effect of capillary fouling on scattering increases monotonically with the intensity scattered by the material within the X-ray beam. Overlapping peaks, often skewed because of sample interaction with the column matrix, can now be accurately decomposed using non-symmetrical modified Gaussian functions. As an example, the case of a polydisperse solution of aldolase is analyzed: from heavily convoluted peaks, individual SAXS profiles of tetramers, octamers and dodecamers are extracted and reliably modeled.
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http://dx.doi.org/10.1107/S1600576716011201DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5045733PMC
October 2016

Simultaneous Identification of Spectral Properties and Sizes of Multiple Particles in Solution with Subnanometer Resolution.

Angew Chem Int Ed Engl 2016 09 27;55(39):11770-4. Epub 2016 Jul 27.

Max-Planck-Institut für Kolloid- und Grenzflächenforschung, Kolloidchemie, Forschungscampus Golm, Am Mühlenberg, 14424, Potsdam, Germany.

We report an unsurpassed solution characterization technique based on analytical ultracentrifugation, which demonstrates exceptional potential for resolving particle sizes in solution with sub-nm resolution. We achieve this improvement in resolution by simultaneously measuring UV/Vis spectra while hydrodynamically separating individual components in the mixture. By equipping an analytical ultracentrifuge with a novel multi-wavelength detector, we are adding a new spectral discovery dimension to traditional hydrodynamic characterization, and amplify the information obtained by orders of magnitude. We demonstrate the power of this technique by characterizing unpurified CdTe nanoparticle samples, avoiding tedious and often impossible purification and fractionation of nanoparticles into apparently monodisperse fractions. With this approach, we have for the first time identified the pure spectral properties and band-gap positions of discrete species present in the CdTe mixture.
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http://dx.doi.org/10.1002/anie.201603844DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5148131PMC
September 2016

Defining the Intrinsically Disordered C-Terminal Domain of SSB Reveals DNA-Mediated Compaction.

J Mol Biol 2016 Jan 18;428(2 Pt A):357-364. Epub 2015 Dec 18.

ISIS Spallation Neutron and Muon Source, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, United Kingdom; School of Biosciences, University of Nottingham, Sutton Bonington Campus, Leicestershire LE12 5RD, United Kingdom; Research Complex at Harwell, Rutherford Appleton Laboratory, Oxfordshire OX11 0FA, United Kingdom. Electronic address:

The bacterial single-stranded DNA (ssDNA) binding protein SSB is a strictly conserved and essential protein involved in diverse functions of DNA metabolism, including replication and repair. SSB comprises a well-characterized tetrameric core of N-terminal oligonucleotide binding OB folds that bind ssDNA and four intrinsically disordered C-terminal domains of unknown structure that interact with partner proteins. The generally accepted, albeit speculative, mechanistic model in the field postulates that binding of ssDNA to the OB core induces the flexible, undefined C-terminal arms to expand outwards encouraging functional interactions with partner proteins. In this structural study, we show that the opposite is true. Combined small-angle scattering with X-rays and neutrons coupled to coarse-grained modeling reveal that the intrinsically disordered C-terminal arms are relatively collapsed around the tetrameric OB core and collapse further upon ssDNA binding. This implies a mechanism of action, in which the disordered C-terminal domain collapse traps the ssDNA and pulls functional partners onto the ssDNA.
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http://dx.doi.org/10.1016/j.jmb.2015.12.007DOI Listing
January 2016

Characterization of size, anisotropy, and density heterogeneity of nanoparticles by sedimentation velocity.

Anal Chem 2014 Aug 18;86(15):7688-95. Epub 2014 Jul 18.

Department of Biochemistry, The University of Texas Health Science Center at San Antonio , 7703 Floyd Curl Drive, San Antonio, Texas 78229-3901, United States.

A critical problem in materials science is the accurate characterization of the size dependent properties of colloidal inorganic nanocrystals. Due to the intrinsic polydispersity present during synthesis, dispersions of such materials exhibit simultaneous heterogeneity in density ρ, molar mass M, and particle diameter d. The density increments ∂ρ/∂d and ∂ρ/∂M of these nanoparticles, if known, can then provide important information about crystal growth and particle size distributions. For most classes of nanocrystals, a mixture of surfactants is added during synthesis to control their shape, size, and optical properties. However, it remains a challenge to accurately determine the amount of passivating ligand bound to the particle surface post synthesis. The presence of the ligand shell hampers an accurate determination of the nanocrystal diameter. Using CdSe and PbS semiconductor nanocrystals, and the ultrastable silver nanoparticle (M4Ag44(p-MBA)30), as model systems, we describe a Custom Grid method implemented in UltraScan-III for the characterization of nanoparticles and macromolecules using sedimentation velocity analytical ultracentrifugation. We show that multiple parametrizations are possible, and that the Custom Grid method can be generalized to provide high resolution composition information for mixtures of solutes that are heterogeneous in two out of three parameters. For such cases, our method can simultaneously resolve arbitrary two-dimensional distributions of hydrodynamic parameters when a third property can be held constant. For example, this method extracts partial specific volume and molar mass from sedimentation velocity data for cases where the anisotropy can be held constant, or provides anisotropy and partial specific volume if the molar mass is known.
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http://dx.doi.org/10.1021/ac501722rDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4144751PMC
August 2014

A parametrically constrained optimization method for fitting sedimentation velocity experiments.

Biophys J 2014 Apr;106(8):1741-50

The University of Texas Health Science Center at San Antonio, Department of Biochemistry, San Antonio, Texas. Electronic address:

A method for fitting sedimentation velocity experiments using whole boundary Lamm equation solutions is presented. The method, termed parametrically constrained spectrum analysis (PCSA), provides an optimized approach for simultaneously modeling heterogeneity in size and anisotropy of macromolecular mixtures. The solutions produced by PCSA are particularly useful for modeling polymerizing systems, where a single-valued relationship exists between the molar mass of the growing polymer chain and its corresponding anisotropy. The PCSA uses functional constraints to identify this relationship, and unlike other multidimensional grid methods, assures that only a single molar mass can be associated with a given anisotropy measurement. A description of the PCSA algorithm is presented, as well as several experimental and simulated examples that illustrate its utility and capabilities. The performance advantages of the PCSA method in comparison to other methods are documented. The method has been added to the UltraScan-III software suite, which is available for free download from http://www.ultrascan.uthscsa.edu.
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http://dx.doi.org/10.1016/j.bpj.2014.02.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4008825PMC
April 2014

Fibrinogen species as resolved by HPLC-SAXS data processing within the () enhanced SAS module.

J Appl Crystallogr 2013 Dec 15;46(Pt 6):1823-1833. Epub 2013 Nov 15.

Department of Biochemistry, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.

Fibrinogen is a large heterogeneous aggregation/degradation-prone protein playing a central role in blood coagulation and associated pathologies, whose structure is not completely resolved. When a high-molecular-weight fraction was analyzed by size-exclusion high-performance liquid chromatography/small-angle X-ray scattering (HPLC-SAXS), several composite peaks were apparent and because of the stickiness of fibrinogen the analysis was complicated by severe capillary fouling. Novel SAS analysis tools developed as a part of the (; http://somo.uthscsa.edu/), an open-source suite of utilities with advanced graphical user interfaces whose initial goal was the hydrodynamic modeling of biomacromolecules, were implemented and applied to this problem. They include the correction of baseline drift due to the accumulation of material on the SAXS capillary walls, and the Gaussian decomposition of non-baseline-resolved HPLC-SAXS elution peaks. It was thus possible to resolve at least two species co-eluting under the fibrinogen main monomer peak, probably resulting from in-column degradation, and two others under an oligomers peak. The overall and cross-sectional radii of gyration, molecular mass and mass/length ratio of all species were determined using the manual or semi-automated procedures available within the SAS module. Differences between monomeric species and linear and sideways oligomers were thus identified and rationalized. This new version additionally contains several computational and graphical tools, implementing functionalities such as the mapping of residues contributing to particular regions of (), and an advanced module for the comparison of primary () data with model curves computed from atomic level structures or bead models. It should be of great help in multi-resolution studies involving hydrodynamics, solution scattering and crystallographic/NMR data.
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http://dx.doi.org/10.1107/S0021889813027751DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3831300PMC
December 2013

Characterization of reversible associations by sedimentation velocity with UltraScan.

Macromol Biosci 2010 Jul;10(7):775-82

Department of Biochemistry, The University of Texas Health Science Center at San Antonio, San Antonio, Texas 78229, USA.

We compare here the utility of sedimentation velocity (SV) to sedimentation equilibrium (SE) analysis for the characterization of reversible systems. Genetic algorithm optimization in UltraScan is used to optimize the model and to obtain solution properties of all components present in the system. We apply our method to synthetic and experimental data, and suggest limits for the accessible kinetic range. We conclude that equilibrium constants obtained from SV and SE analysis are equivalent, but that SV experiments provide better confidence for the K(d), can better account for the presence of contaminants and provide additional information including rate constants and shape parameters.
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http://dx.doi.org/10.1002/mabi.200900481DOI Listing
July 2010

Developments in the US-SOMO bead modeling suite: new features in the direct residue-to-bead method, improved grid routines, and influence of accessible surface area screening.

Macromol Biosci 2010 Jul;10(7):746-53

Department of Biochemistry, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, TX 78229, USA.

The US-SOMO suite provides a flexible interface for accurately computing solution parameters from 3D structures of biomacromolecules through bead-modeling approaches. We present an extended analysis of the influence of accessible surface area screening, overlap reduction routines, and approximations for non-coded residues and missing atoms on the computed parameters for models built by the residue-to-bead direct correspondence and the cubic grid methods. Importantly, by taking the theoretical hydration into account at the atomic level, the performance of the grid-type models becomes comparable or exceeds that of the corresponding hydrated residue-to-bead models.
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http://dx.doi.org/10.1002/mabi.200900474DOI Listing
July 2010

The Open AUC Project.

Eur Biophys J 2010 Feb 19;39(3):347-59. Epub 2009 Mar 19.

Colloid Chemistry, Max-Planck-Institute of Colloids and Interfaces, Research Campus Golm, Am Mühlenberg, 14424 Potsdam, Germany.

Progress in analytical ultracentrifugation (AUC) has been hindered by obstructions to hardware innovation and by software incompatibility. In this paper, we announce and outline the Open AUC Project. The goals of the Open AUC Project are to stimulate AUC innovation by improving instrumentation, detectors, acquisition and analysis software, and collaborative tools. These improvements are needed for the next generation of AUC-based research. The Open AUC Project combines on-going work from several different groups. A new base instrument is described, one that is designed from the ground up to be an analytical ultracentrifuge. This machine offers an open architecture, hardware standards, and application programming interfaces for detector developers. All software will use the GNU Public License to assure that intellectual property is available in open source format. The Open AUC strategy facilitates collaborations, encourages sharing, and eliminates the chronic impediments that have plagued AUC innovation for the last 20 years. This ultracentrifuge will be equipped with multiple and interchangeable optical tracks so that state-of-the-art electronics and improved detectors will be available for a variety of optical systems. The instrument will be complemented by a new rotor, enhanced data acquisition and analysis software, as well as collaboration software. Described here are the instrument, the modular software components, and a standardized database that will encourage and ease integration of data analysis and interpretation software.
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http://dx.doi.org/10.1007/s00249-009-0438-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2812709PMC
February 2010

A two-dimensional spectrum analysis for sedimentation velocity experiments of mixtures with heterogeneity in molecular weight and shape.

Eur Biophys J 2010 Feb 27;39(3):405-14. Epub 2009 Feb 27.

Department of Biochemistry, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, MC 7760, San Antonio, TX 78229-3901, USA.

We report a model-independent analysis approach for fitting sedimentation velocity data which permits simultaneous determination of shape and molecular weight distributions for mono- and polydisperse solutions of macromolecules. Our approach allows for heterogeneity in the frictional domain, providing a more faithful description of the experimental data for cases where frictional ratios are not identical for all components. Because of increased accuracy in the frictional properties of each component, our method also provides more reliable molecular weight distributions in the general case. The method is based on a fine grained two-dimensional grid search over s and f/f (0), where the grid is a linear combination of whole boundary models represented by finite element solutions of the Lamm equation with sedimentation and diffusion parameters corresponding to the grid points. A Monte Carlo approach is used to characterize confidence limits for the determined solutes. Computational algorithms addressing the very large memory needs for a fine grained search are discussed. The method is suitable for globally fitting multi-speed experiments, and constraints based on prior knowledge about the experimental system can be imposed. Time- and radially invariant noise can be eliminated. Serial and parallel implementations of the method are presented. We demonstrate with simulated and experimental data of known composition that our method provides superior accuracy and lower variance fits to experimental data compared to other methods in use today, and show that it can be used to identify modes of aggregation and slow polymerization.
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http://dx.doi.org/10.1007/s00249-009-0413-5DOI Listing
February 2010

The implementation of SOMO (SOlution MOdeller) in the UltraScan analytical ultracentrifugation data analysis suite: enhanced capabilities allow the reliable hydrodynamic modeling of virtually any kind of biomacromolecule.

Eur Biophys J 2010 Feb 21;39(3):423-35. Epub 2009 Feb 21.

Department of Biochemistry, The University of Texas Health Science Center at San Antonio, San Antonio, TX, USA.

The interpretation of solution hydrodynamic data in terms of macromolecular structural parameters is not a straightforward task. Over the years, several approaches have been developed to cope with this problem, the most widely used being bead modeling in various flavors. We report here the implementation of the SOMO (SOlution MOdeller; Rai et al. in Structure 13:723-734, 2005) bead modeling suite within one of the most widely used analytical ultracentrifugation data analysis software packages, UltraScan (Demeler in Modern analytical ultracentrifugation: techniques and methods, Royal Society of Chemistry, UK, 2005). The US-SOMO version is now under complete graphical interface control, and has been freed from several constraints present in the original implementation. In the direct beads-per-atoms method, virtually any kind of residue as defined in the Protein Data Bank (e.g., proteins, nucleic acids, carbohydrates, prosthetic groups, detergents, etc.) can be now represented with beads whose number, size and position are all defined in user-editable tables. For large structures, a cubic grid method based on the original AtoB program (Byron in Biophys J 72:408-415, 1997) can be applied either directly on the atomic structure, or on a previously generated bead model. The hydrodynamic parameters are then computed in the rigid-body approximation. An extensive set of tests was conducted to further validate the method, and the results are presented here. Owing to its accuracy, speed, and versatility, US-SOMO should allow to fully take advantage of the potential of solution hydrodynamics as a complement to higher resolution techniques in biomacromolecular modeling.
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http://dx.doi.org/10.1007/s00249-009-0418-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2872189PMC
February 2010

Analysis of heterogeneity in molecular weight and shape by analytical ultracentrifugation using parallel distributed computing.

Methods Enzymol 2009 ;454:87-113

Department of Biochemistry, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, USA.

A computational approach for fitting sedimentation velocity experiments from an analytical ultracentrifuge in a model-independent fashion is presented. This chapter offers a recipe for obtaining high-resolution information for both the shape and the molecular weight distributions of complex mixtures that are heterogeneous in shape and molecular weight and provides suggestions for experimental design to optimize information content. A combination of three methods is used to find the solution most parsimonious in parameters and to verify the statistical confidence intervals of the determined parameters. A supercomputer implementation with a MySQL database back end is integrated into the UltraScan analysis software. The UltraScan LIMS Web portal is used to perform the calculations through a Web interface. The performance and limitations of the method when employed for the analysis of complex mixtures are demonstrated using both simulated data and experimental data characterizing amyloid aggregation.
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http://dx.doi.org/10.1016/S0076-6879(08)03804-4DOI Listing
April 2009