Publications (127)169.9 Total impact
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ABSTRACT: Cryo EM structures of maturationintermediate Prohead I of bacteriophage HK97 with (PhI(Pro+)) and without (PhI(Pro)) the viral protease packaged have been reported (Veesler et al., Structure 22:230, 2014). In spite of PhI(Pro+) containing an additional ∼100×24kD of protein, the two structures appeared identical although the two particles have substantially different biochemical properties, e.g., PhI(Pro) is less stable to disassembly conditions such as urea. Here the same cryo EM images are used to characterize the spatial heterogeneity of the particles at 17Å resolution by variance analysis and show that PhI(Pro) has roughly twice the standard deviation of PhI(Pro+). Furthermore, the greatest differences in standard deviation are present in the region where the δdomain, not seen in xray crystallographic structures or fully seen in cryo EM, is expected to be located. Thus presence of the protease appears to stabilize the δdomain which the protease will eventually digest.  [Show abstract] [Hide abstract]
ABSTRACT: To describe a toolkit of components for mathematical models of the relationship between cortical neural activity and spaceresolved and timeresolved flows and volumes of oxygenated and deoxygenated hemoglobin motivated by optical intrinsic signal imaging (OISI). Both blood flow and blood volume and both oxygenated and deoxygenated hemoglobin and their interconversion are accounted for. Flow and volume are described by including analogies to both resistive and capacitive electrical circuit elements. Oxygenated and deoxygenated hemoglobin and their interconversion are described by generalization of Kirchhoff's laws based on wellmixed compartments. Mathematical models built from this toolkit are able to reproduce experimental singlestimulus OISI results that are described in papers from other research groups and are able to describe the response to multiplestimuli experiments as a sublinear superposition of responses to the individual stimuli. The same assembly of tools from the toolkit but with different parameter values is able to describe effects that are considered distinctive, such as the presence or absence of an initial decrease in oxygenated hemoglobin concentration, indicating that the differences might be due to unique parameter values in a subject rather than different fundamental mechanisms.  [Show abstract] [Hide abstract]
ABSTRACT: Restingstate functional MRI (rs fMRI) is widely used to noninvasively study human brain networks. Network functional connectivity is estimated by calculating the standard correlation between bloodoxygenlevel dependent (BOLD) signals in specific regions of interests (ROIs). However, standard correlation fails to characterize the causality and the direction of information flow between regions, which are important factors in characterizing a network. Here, we use causal linear timeinvariant models, with the impulse response duration estimated by Information Criteria, to describe the effective connectivity between ROIs. To do so, we replace the standard correlation between BOLD signals with a correlation between a BOLD signal and a prediction via the model of that BOLD signal. Prediction correlation is then used in a network analysis similar to that used with standard correlation. Our results include the causality information, the direction of information flow, and the possibility of delays in information flow. This approach replicates the local and distributed network architecture of the human brain previously observed with standard correlations, as well as providing novel insight into the directed interactivity of regions comprising these networks.  [Show abstract] [Hide abstract]
ABSTRACT: Quasiequivalent viruses that infect animals and bacteria require a maturation process in which particles transition from initially assembled procapsids to infectious virions. Nudaurelia capensis ω virus (NωV) is a T = 4, eukaryotic, singlestranded ribonucleic acid virus that has proved to be an excellent model system for studying the mechanisms of viral maturation. Structures of NωV procapsids (diameter = 480 Å), a maturation intermediate (410 Å), and the mature virion (410 Å) were determined by electron cryomicroscopy and threedimensional image reconstruction (cryoEM). The cryoEM density for each particle type was analyzed with a recently developed maximum likelihood variance (MLV) method for characterizing microstates occupied in the ensemble of particles used for the reconstructions. The procapsid and the mature capsid had overall low variance (i.e., uniform particle populations) while the maturation intermediate (that had not undergone postassembly autocatalytic cleavage) had roughly two to four times the variance of the first two particles. Without maturation cleavage, the particles assume a variety of microstates, as the frustrated subunits cannot reach a minimum energy configuration. Geometric analyses of subunit coordinates provided a quantitative description of the particle reorganization during maturation. Superposition of the four quasiequivalent subunits in the procapsid had an average root mean square deviation (RMSD) of 3 Å while the mature particle had an RMSD of 11 Å, showing that the subunits differentiate from near equivalent environments in the procapsid to strikingly nonequivalent environments during maturation. Autocatalytic cleavage is clearly required for the reorganized mature particle to reach the minimum energy state required for stability and infectivity. Copyright © 2014 John Wiley & Sons, Ltd. 
Conference Paper: 3D Statistical Characterization of the Heterogeneity of Biological Macromolecular Complexes by Electron Microscopy
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ABSTRACT: Electron microscopy provides images of macromolecular complexes from which the 3D structure of the complex can be computed when all instances of a complex are identical. An algorithm for characterizing the 3D spatial statistics of the complex is described and demonstrated for the important case when different instances are not identical.  [Show abstract] [Hide abstract]
ABSTRACT: We generalize the concept of allostery from the traditional non activesite control of enzymes to virus maturation. Virtually all animal viruses transition from a procapsid, non infectious state, to a mature infectious state. The procapsid contains an encoded chemical program that is executed following an environmental cue. We developed an exceptionally accessible virus system for the study of the activators of maturation and the down stream consequences that result in particle stability and infectivity.Nudaurelia Capensis Omegavirus (NωV) is a T=4 icosahedral virus that undergoes a dramatic maturation in which the 490Å spherical procapsid condenses to a 400Å icosahedral shaped capsid with associated specific autoproteolysis and stabilization. Employing Xray crystallography, time resolved electron cryomicroscopy and hydrogen/deuterium exchange as well as biochemistry it was possible to define the mechanisms of allosteric communication among the 4 quasiequivalent subunits in the icosahedral asymmetric unit. These gene products undergo proteolysis at different rates, dependent on quaternary structure environment, while particle stability is conferred globally following only a few local subunit transitions. We show that there is a close similarity between the concepts of tensegrity, associated with geodesic domes and mechanical engineering, and allostery, associated with biochemical control mechanisms. 
Article: Dynamics in Cryo EM Reconstructions Visualized with MaximumLikelihood Derived Variance Maps.
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ABSTRACT: CryoEM data capture the dynamic character associated with biological macromolecular assemblies by preserving the various conformations of the individual specimens at the moment of flash freezing. Regions of high variation in the data set are apparent in the image reconstruction due to the poor density that results from the lack of superposition of these regions. These observations are qualitative and, to date, only preliminary efforts have been made to quantitate the heterogeneity in the ensemble of particles that are individually imaged. We developed and tested a quantitative method for simultaneously computing a reconstruction of the particle and a map of the spacevarying heterogeneity of the particle based on an entire data set. The method uses a maximum likelihood algorithm that explicitly takes into account the continuous variability from one instance to another instance of the particle. The result describes the heterogeneity of the particle as a variance to be plotted at every voxel of the reconstructed density. The test, employing time resolved data sets of virus maturation, not only recapitulated local variations obtained with difference map analysis, but revealed a remarkable time dependent reduction in the overall particle dynamics that was unobservable with classical methods of analysis. 
Conference Paper: Describing the structure of a macro molecular complex as a random Signal in noise and a maximum likelihood reconstruction
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ABSTRACT: Instances of biological macromolecular complexes that have identical chemical constituents may not have the same geometry due to, for example, flexibility. Cryo electron microscopy provides one noisy projection image of each of many instances of a complex where the projection directions for the different instances are random. The noise is sufficient severe (SNR ≪ 1) that the projection direction for a particular image cannot be easily estimated from the individual image. The goal is to determine the 3D geometry of the complex (the 3D distribution of electron scattering intensity) which requires fusing information from these many images of many complexes. In order to describe the geometric heterogeneity of the complexes, the complex is described as a weighted sum of basis functions where the weights are random. In order to get tractable algorithms, the weights are modeled as Gaussian random variables with unknown statistics and the noise is modeled as additive Gaussian random variables with unknown covariance. The statistics of the weights and the statistics of the noise are jointly estimated by maximum likelihood by a generalized expectation maximization algorithm. The method has been developed to the point where it appears to be able to solve problems of interest to the structural biology community.  [Show abstract] [Hide abstract]
ABSTRACT: An estimation problem for statistical reconstruction of heterogeneous threedimensional objects from twodimensional tomographic data (singleparticle cryoelectron microscope images) is posed as the problem of estimating class probabilities, means, and covariances for a Gaussian mixture where both the mean and covariance are stochastically structured. Both discrete (i.e., classes) and continuous heterogeneity is included. A maximum likelihood solution computed by a generalized expectationmaximization algorithm is presented and demonstrated on experimental images of Flock House Virus.  [Show abstract] [Hide abstract]
ABSTRACT: The instantaneous rate of change of alcohol exposure (slope) may contribute to changes in measures of brain function following administration of alcohol that are usually attributed to breath alcohol concentration (BrAC) acting alone. To test this proposition, a 2session experiment was designed in which carefully prescribed, constantslope trajectories of BrAC intersected at the same exposure level and time since the exposure began. This paper presents the methods and limitations of the experimental design. Individualized intravenous infusion rate profiles of 6% ethanol (EtOH) that achieved the constantslope trajectories for an individual were precomputed using a physiologically based pharmacokinetic model. Adjusting the parameters of the model allowed each infusion profile to account for the subject's EtOH distribution and elimination kinetics. Sessions were conducted in randomized order and made no use of feedback of BrAC measurements obtained during the session to modify the precalculated infusion profiles. In one session, an individual's time course of exposure, BrAC(t), was prescribed to rise at a constant rate of 6.0 mg% per minute until it reached 68 mg% and then descend at 1.0 mg% per minute; in the other, to rise at a rate of 3.0 mg% per minute. The 2 exposure trajectories were designed to intersect at a BrAC (t = 20 minutes) = 60 mg% at an experimental time of 20 minutes. Intersection points for 54 of 61 subjects were within prescribed deviations (range of ± 3 mg% and ± 4 minutes from the nominal intersection point). Results confirmed the feasibility of applying the novel methods for achieving the intended time courses of the BrAC, with technical problems limiting success to 90% of the individuals tested.  [Show abstract] [Hide abstract]
ABSTRACT: Instances of biological macromolecular complexes that have identical chemical constituents may not have the same geometry due to, for example, flexibility. Cryo electron microscopy provides one noisy projection image of each of many instances of a complex where the projection directions for the different instances are random. The noise is sufficient severe (SNR << 1) that the projection direction for a particular image cannot be easily estimated from the individual image. The goal is to determine the 3D geometry of the complex (the 3D distribution of electron scattering intensity) which requires fusing information from these many images of many complexes. In order to describe the geometric heterogeneity of the complexes, the complex is described as a weighted sum of basis functions where the weights are random. In order to get tractable algorithms, the weights are modeled as Gaussian random variables with unknown statistics and the noise is modeled as additive Gaussian random variables with unknown covariance. The statistics of the weights and the statistics of the noise are jointly estimated by maximum likelihood by a generalized expectation maximization algorithm. The method has been developed to the point where it appears able to solve problems of interest to the structural biology community.  [Show abstract] [Hide abstract]
ABSTRACT: A highly scalable method for determining the projection orientation of each image in a set of cryo electron microscopy images of a labeled particle is proposed. The method relies on the presence of a label that is a sufficiently strong scatterer such that its 2D location in each image can be restricted to at most a small number of sites by processing applied to each image individually. It is not necessary to know the 3D location of the label on the particle. After first determining the possible locations of the label in the 2D images in parallel, the information from all images is fused to determine the 3D location of the label on the particle and then the 3D location is used to determine the projection orientation for each image by processing each image individually. With projection orientations, many algorithms exist for computing the 3D reconstruction. The performance of the algorithm is studied as a function of the label SNR.  [Show abstract] [Hide abstract]
ABSTRACT: Subtle alterations in cerebral blood flow can impact the health and function of brain cells and are linked to cognitive decline and dementia. To understand hemodynamics in the threedimensional vascular network of the cerebral cortex, we applied twophoton excited fluorescence microscopy to measure the motion of red blood cells (RBCs) in individual microvessels throughout the vascular hierarchy in anesthetized mice. To resolve heartbeat and respirationdependent flow dynamics, we simultaneously recorded the electrocardiogram and respiratory waveform. We found that centerline RBC speed decreased with decreasing vessel diameter in arterioles, slowed further through the capillary bed, and then increased with increasing vessel diameter in venules. RBC flow was pulsatile in nearly all cortical vessels, including capillaries and venules. Heartbeatinduced speed modulation decreased through the vascular network, while the delay between heartbeat and the time of maximum speed increased. Capillary tube hematocrit was 0.21 and did not vary with centerline RBC speed or topological position. Spatial RBC flow profiles in surface vessels were blunted compared with a parabola and could be measured at vascular junctions. Finally, we observed a transient decrease in RBC speed in surface vessels before inspiration. In conclusion, we developed an approach to study detailed characteristics of RBC flow in the threedimensional cortical vasculature, including quantification of fluctuations in centerline RBC speed due to cardiac and respiratory rhythms and flow profile measurements. These methods and the quantitative data on basal cerebral hemodynamics open the door to studies of the normal and diseasedstate cerebral microcirculation. 
Conference Paper: Measurements of Pulsatile Hemodynamics In Brain Microvessels
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ABSTRACT: Experimental measurements are reported of blood flow in the microcirculation of the brain cortex. We use twophoton excited fluorescence (2PEF) microscopy to measure cortical blood flow in anesthetized rats. 2PEF is a nonlinear optical technique that allows detection of fluorescence from a point in highly scattering samples such as neural tissue. By rapidly scanning the detection point throughout the sample, an “image” can be detected deep inside the cortex of a live animal. We exploit this technique to track the motion of individual red blood cells inside arterioles, capillaries, and venules in the cortex with precision that is unmatched by other in vivo measurement methods. Furthermore, we correlate the blood cell speed with heartbeat and respiration measurements to extract detailed information about timedependent blood flow in individual vessels and at vessel bifurcations. Measurements are made in surface vessels and in capillaries deep in the brain tissue. Timeaveraged blood flow speed decreases with vessel diameter in arterioles and in capillaries, and increases in venules. Blood flow speed decreases with inspiration by about 20% across all parts of the cardiac cycle. These results show that hemodynamics in the brain cortex can be resolved with high spatial and temporal resolution in vivo, which is a critical step toward identifying and quantifying cerebral blood flow abnormalities associated with a variety of hematological disorders.  [Show abstract] [Hide abstract]
ABSTRACT: The influence of family history and genetics on the risk for the development of abuse or dependence is a major theme in alcoholism research. Recent research have used endophenotypes and behavioral paradigms to help detect further genetic contributions to this disease. Electronic tasks, essentially video games, which provide alcohol as a reward in controlled environments and with specified exposures have been developed to explore some of the behavioral and subjective characteristics of individuals with or at risk for alcohol substance use disorders. A generative model (containing parameters with unknown values) of a simple game involving a progressive work paradigm is described along with the associated point process signal processing that allows system identification of the model. The system is demonstrated on human subject data. The same human subject completing the task under different circumstances, e.g., with larger and smaller alcohol reward values, is assigned different parameter values. Potential meanings of the different parameter values are described.  [Show abstract] [Hide abstract]
ABSTRACT: The cortical microvasculature plays a key role in cortical tissue health by transporting important molecules via blood. Disruptions to blood flow in the microvasculature due to events such as stroke can thus induce damage to the cortex. Recent developments in twophoton microscopy have enabled in vivo imaging of anesthetized rat cortex in three dimensions. The microscopy data provide information about the geometry of the cortical microvasculature, length and diameter of the vessels in the imaged microvasculature network, and blood flow through a subset of those vessels. We demonstrate a model that achieves three goals. First, given a network of interconnected vessels and flow measurements on a subset of those vessels, we can estimate the flows in the remaining vessels. Second, we can determine which and how many vessels should have blood flow measurements taken to provide sufficient information to predict the unmeasured flows. Finally, the model enables us to predict effects of blockages in one or more vessels, indicating which vessels are most important to overall flow in the network.  [Show abstract] [Hide abstract]
ABSTRACT: Many micro to nanoscale 3D biological objects have a helical symmetry. Cryo electron microscopy provides 2D projection images where, however, the images have low SNR and unknown projection directions. The object is described as a helical array of identical motifs, where both the parameters of the helical symmetry and the motif are unknown. Using a detailed image formation model, a maximumlikelihood estimator for the parameters of the symmetry and the 3D motif based on images of many objects and algorithms for computing the estimate are described. The possibility that the objects are not identical but rather come from a small set of homogeneous classes is included. The first example is based on 316 128 ×100 pixel experimental images of Tobacco Mosaic Virus, has one class, and achieves 12.40Å spatial resolution in the reconstruction. The second example is based on 400 128 ×128 pixel synthetic images of helical objects constructed from NaK ion channel pore macromolecular complexes, has two classes differing in helical symmetry, and achieves 7.84 and 7.90Å spatial resolution in the reconstructions for the two classes. 
Conference Paper: Understanding dynamics of biological macromolecular complexes by estimating a mechanical model via statistical mechanics from cryo electron microscopy images
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ABSTRACT: Cryo electron microscopy (cryo EM) imaging experiments can lead to stochastic models for biological macromolecular complexes. However, interpreting the statistical variability is difficult. In some situations, the variability in the original complexes is due primarily to thermal fluctuations which are snap frozen in place during the preparation of the specimen. In this case the images are images of samples of the equilibrium statistical mechanics ensemble of the complex. Based on representing the complex by a springmass mechanical model, an estimation problem for determining the masses and spring constants is described and demonstrated on synthetic data. With a model, quantities such as normal modes can be computed, which provide insight into the dynamics of biological complexes. 
Conference Paper: Stochastic 3D signal reconstruction from noisy projection data for heterogeneous instances of objects in electron microscopy imagery
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ABSTRACT: Different instances of a biological macromolecular complex need not share the same 3D electron scattering distribution function due to stoichiometric variability, flexibility and vibrations, etc. Cryo electron microscopy provides 2D images of each of many such complexes where each image is roughly a projection. A statistical estimation problem is described and demonstrated for determining a statistical description of the complex from such imagery.  [Show abstract] [Hide abstract]
ABSTRACT: Sulfolobus Turreted Icosahedral Virus (STIV) experiences an extracellular environment of near boiling acid (80°C, pH 3) and particles purified under these conditions were previously analyzed by cryo electron microscopy and image reconstruction. Here we describe cryotomograms of Solfolobus cells infected with STIV and the maximum likelihood algorithm employed to compute reconstructions of virions within the cell. Virions in four different tomograms were independently reconstructed with an average of 91 particles per tomogram and their structures compared with each other and with the higher resolution singleparticle reconstruction from purified virions. The algorithm described here automatically classified and oriented two different particle types within each cell and generated reconstructions of full and empty particles. Because the particles are randomly oriented within the cell, the reconstructions do not suffer from the missing wedge of data absent from the reciprocalspace tomogram. The fact that the particles have icosahedral symmetry is used to dramatically improve the signal to noise ratio in the reconstructions. The reconstructions have approximately 60Å resolution (based on Fourier Shell Correlation analysis among reconstructions computed by the algorithm described here from four different tomograms).
Publication Stats
736  Citations  
169.90  Total Impact Points  
Top Journals
Institutions

20092015

Cornell University
 Department of Biomedical Engineering
Итак, New York, United States


19912011

Purdue University
 School of Electrical and Computer Engineering
ウェストラファイエット, Indiana, United States


2005

The Scripps Research Institute
 Department of Cell and Molecular Biology
لا هویا, California, United States


1997

University of Louisiana at Lafayette
 Department of Electrical & Computer Engineering
Lafayette, Louisiana, United States
