In this article we present a method to identify and extract spherical particle images from noisy ... more In this article we present a method to identify and extract spherical particle images from noisy spot-scan and flood beam electron microscopy images. We use a template matching algorithm with additional image preprocessing operations to allow consistent particle selection in spot-scan and other highly spatially varying images. In addition, this algorithm incorporates an automated image-cutting and edge-sewing mechanism that allows
Image formation in bright field electron microscopy can be described with the help of the contras... more Image formation in bright field electron microscopy can be described with the help of the contrast transfer function (CTF). In this work the authors describe the "CTF Estimation Challenge", called by the Madrid Instruct Image Processing Center (I2PC) in collaboration with the National Center for Macromolecular Imaging (NCMI) at Houston. Correcting for the effects of the CTF requires accurate knowledge of the CTF parameters, but these have often been difficult to determine. In this challenge, researchers have had the opportunity to test their ability in estimating some of the key parameters of the electron microscope CTF on a large micrograph data set produced by well-known laboratories on a wide set of experimental conditions. This work presents the first analysis of the results of the CTF Estimation Challenge, including an assessment of the performance of the different software packages under different conditions, so as to identify those areas of research where further developments would be desirable in order to achieve high-resolution structural information.
Symposium on geometry processing : [proceedings] / sponsored by the Eurographics Association in cooperation with ACM SIGGRAPH. Symposium on Geometry Processing, 2003
As the logical next step after sequencing the mouse genome, biologists have developed laboratory ... more As the logical next step after sequencing the mouse genome, biologists have developed laboratory methods for rapidly determining where each of the 30K genes in the mouse genome is synthesizing protein. Applying these methods to the mouse brain, biologists are currently generating large numbers of 2D cross-sectional images that record the expression pattern for each gene in the mouse genome. In this paper, we describe the structure of a geometric database for the mouse brain that allows biologists to organize and search this gene expression data. The central component of this database is an atlas that explicitly partitions the mouse brain into key anatomical regions. This atlas is represented as a Catmull-Clark subdivision mesh with anatomical regions separated by a network of B-spline crease curves. New gene expression images are added to the database by deforming this atlas onto each image using techniques developed for fitting subdivision surfaces to scatter data. Due to this part...
The three-dimensional structure of rice dwarf virus was determined to 6.8 A resolution by single ... more The three-dimensional structure of rice dwarf virus was determined to 6.8 A resolution by single particle electron cryomicroscopy. By integrating the structural analysis with bioinformatics, the folds of the proteins in the double-shelled capsid were derived. In the outer shell protein, the uniquely orientated upper and lower domains are composed of similar secondary structure elements but have different relative orientations from that of bluetongue virus in the same Reoviridae family. Differences in both sequence and structure between these proteins may be important in defining virus-host interactions. The inner shell protein adopts a conformation similar to other members of Reoviridae, suggesting a common ancestor that has evolved to infect hosts ranging from plants to animals. Symmetry mismatch between the two shells results in nonequivalent, yet specific, interactions that contribute to the stability of this large macromolecular machine.
Herpes simplex virus type 1 virions were examined by electron cryomicroscopy, allowing the three-... more Herpes simplex virus type 1 virions were examined by electron cryomicroscopy, allowing the three-dimensional structure of the infectious particle to be visualized for the first time. The capsid shell is identical to that of B-capsids purified from the host cell nucleus, with the exception of the penton channel, which is closed. The double-stranded DNA genome is organized as regularly spaced ( approximately 26 A) concentric layers inside the capsid. This pattern suggests a spool model for DNA packaging, similar to that for some bacteriophages. The bulk of the tegument is not icosahedrally ordered. However, a small portion appears as filamentous structures around the pentons, interacting extensively with the capsid. Their locations and interactions suggest possible roles for the tegument proteins in regulating DNA transport through the penton channel and binding to cellular transport proteins during viral infection.
Electron energy loss spectroscopy and dark-field imaging in a scanning transmission electron micr... more Electron energy loss spectroscopy and dark-field imaging in a scanning transmission electron microscope were used to determine the thickness of glucose-embedded crotoxin complex crystals. The results demonstrate the feasibility of identifying protein crystals with a thickness of half a unit cell (12.8 nm) under low-dose and low-temperature conditions. The accuracy of this method is limited by the amount of surface coating of the crystal's embedding glucose used for preserving the high-resolution structure of the protein. The histogram of the crystal thickness distribution and the spread of the anticipated crystal thickness allow us to make an estimate of the uncertainty in the glucose layer thickness. This approach can be incorporated as part of the experimental procedure in the three-dimensional data collection for structure determination of protein crystals with variable thicknesses. The measurement can be done on areas approximately 200 nm in diameter so that crystals of suit...
Single-particle analysis has become an increasingly important method for structural determination... more Single-particle analysis has become an increasingly important method for structural determination of large macromolecular assemblies. GroEL is an 800 kDa molecular chaperone, which, along with its co-chaperonin GroES, promotes protein folding both in ...
Background: Bluetongue virus (BTV), which belongs to the Reoviridae family and orbivirus genus, i... more Background: Bluetongue virus (BTV), which belongs to the Reoviridae family and orbivirus genus, is a non-enveloped, icosahedral, double-stranded RNA virus. Several protein layers enclose its genome; upon cell entry the outer layer is stripped away leaving a core, the surface of which is composed of VP7. The structure of the trimeric VP7 molecule has previously been determined using X-ray crystallography.
This paper presents a weight function technique for calculating the stress intensity factors for ... more This paper presents a weight function technique for calculating the stress intensity factors for composite repairs to cracks emanating from an internal notch, corrosion blend out, or a free edge under arbitrary loading in rib stiffened panels. The predictions are compared with both finite element and experimental values. This methodology represents a significant extension to existing assessment and design formulae
A three-dimensional reconstruction of a thin protein crystal requires an accurate assignment of a... more A three-dimensional reconstruction of a thin protein crystal requires an accurate assignment of amplitudes and phases in the three-dimensional reciprocal space. This assignment depends upon the tilt parameters which are experimentally determined. A theoretical estimate is made of the tolerable error in the determination of tilt parameters as a function of resolution and crystal thickness.
... ULTRASTRUCTURE RESEARCH 79, 285-293 (1982)Characterization of Crystalline Filtrate Tetanus To... more ... ULTRASTRUCTURE RESEARCH 79, 285-293 (1982)Characterization of Crystalline Filtrate Tetanus ToxinWAH CHIU,*'1 DEBBY RANKERT,* MARY A. CUMMING,t ... We thank Dr. R. Salemme for his discussion.This work has been supported by US Public HealthService Grants ...
Co-chaperonin protein 10 (cpn10, GroES in Escherichia coli) is a ring-shaped heptameric protein t... more Co-chaperonin protein 10 (cpn10, GroES in Escherichia coli) is a ring-shaped heptameric protein that facilitates substrate folding when in complex with cpn60 (GroEL in E. coli). The cpn10 from the hyperthermophilic, ancient bacterium Aquifex aeolicus (Aacpn10) has a 25-residue C-terminal extension in each monomer not found in any other cpn10 protein. Earlier in vitro work has shown that this tail
IN double-stranded-RNA (dsRNA) viruses found in animals1, bacteria2 and yeast3, the genome is tra... more IN double-stranded-RNA (dsRNA) viruses found in animals1, bacteria2 and yeast3, the genome is transcribed within the structurally intact core of the virion with extraordinary efficiency. The structural organization of the genome and the enzymes involved in the transcription inside any of these viruses, critical for understanding this process, is not known. Here we report what we believe is the first
Massive amounts of data are being generated in an effort to represent for the brain the expressio... more Massive amounts of data are being generated in an effort to represent for the brain the expression of all genes at cellular resolution. Critical to exploiting this effort is the ability to place these data into a common frame of reference. Here we have developed a computational method for annotating gene expression patterns in the context of a digital atlas
Proceedings of the National Academy of Sciences of the United States of America, Jan 9, 2015
Lemon-shaped viruses are common in nature but so far have been observed to infect only archaea. D... more Lemon-shaped viruses are common in nature but so far have been observed to infect only archaea. Due to their unusual shape, the structures of these viruses are challenging to study and therefore poorly characterized. Here, we have studied haloarchaeal virus His1 using cryo-electron tomography as well as biochemical dissociation. The virions have different sizes, but prove to be extremely stable under various biochemical treatments. Subtomogram averaging of the computationally extracted virions resolved a tail-like structure with a central tail hub density and six tail spikes. Inside the tail there are two cavities and a plug density that separates the tail hub from the interior genome. His1 most likely uses the tail spikes to anchor to host cells and the tail hub to eject the genome, analogous to classic tailed bacteriophages. Upon biochemical treatment that releases the genome, the lemon-shaped virion transforms into an empty tube. Such a dramatic transformation demonstrates that t...
A program to implement cross-borehole tomography using a simultaneous iterative reconstruction te... more A program to implement cross-borehole tomography using a simultaneous iterative reconstruction technique (SIRT) has been written. The effectiveness of the program in detecting damages in flat plates has been tested through utilising a combination of computer simulations and experimental tests. Images of sample areas were reconstructed from the generation of simulated data for sample areas containing damage zones of different
In this article we present a method to identify and extract spherical particle images from noisy ... more In this article we present a method to identify and extract spherical particle images from noisy spot-scan and flood beam electron microscopy images. We use a template matching algorithm with additional image preprocessing operations to allow consistent particle selection in spot-scan and other highly spatially varying images. In addition, this algorithm incorporates an automated image-cutting and edge-sewing mechanism that allows
Image formation in bright field electron microscopy can be described with the help of the contras... more Image formation in bright field electron microscopy can be described with the help of the contrast transfer function (CTF). In this work the authors describe the "CTF Estimation Challenge", called by the Madrid Instruct Image Processing Center (I2PC) in collaboration with the National Center for Macromolecular Imaging (NCMI) at Houston. Correcting for the effects of the CTF requires accurate knowledge of the CTF parameters, but these have often been difficult to determine. In this challenge, researchers have had the opportunity to test their ability in estimating some of the key parameters of the electron microscope CTF on a large micrograph data set produced by well-known laboratories on a wide set of experimental conditions. This work presents the first analysis of the results of the CTF Estimation Challenge, including an assessment of the performance of the different software packages under different conditions, so as to identify those areas of research where further developments would be desirable in order to achieve high-resolution structural information.
Symposium on geometry processing : [proceedings] / sponsored by the Eurographics Association in cooperation with ACM SIGGRAPH. Symposium on Geometry Processing, 2003
As the logical next step after sequencing the mouse genome, biologists have developed laboratory ... more As the logical next step after sequencing the mouse genome, biologists have developed laboratory methods for rapidly determining where each of the 30K genes in the mouse genome is synthesizing protein. Applying these methods to the mouse brain, biologists are currently generating large numbers of 2D cross-sectional images that record the expression pattern for each gene in the mouse genome. In this paper, we describe the structure of a geometric database for the mouse brain that allows biologists to organize and search this gene expression data. The central component of this database is an atlas that explicitly partitions the mouse brain into key anatomical regions. This atlas is represented as a Catmull-Clark subdivision mesh with anatomical regions separated by a network of B-spline crease curves. New gene expression images are added to the database by deforming this atlas onto each image using techniques developed for fitting subdivision surfaces to scatter data. Due to this part...
The three-dimensional structure of rice dwarf virus was determined to 6.8 A resolution by single ... more The three-dimensional structure of rice dwarf virus was determined to 6.8 A resolution by single particle electron cryomicroscopy. By integrating the structural analysis with bioinformatics, the folds of the proteins in the double-shelled capsid were derived. In the outer shell protein, the uniquely orientated upper and lower domains are composed of similar secondary structure elements but have different relative orientations from that of bluetongue virus in the same Reoviridae family. Differences in both sequence and structure between these proteins may be important in defining virus-host interactions. The inner shell protein adopts a conformation similar to other members of Reoviridae, suggesting a common ancestor that has evolved to infect hosts ranging from plants to animals. Symmetry mismatch between the two shells results in nonequivalent, yet specific, interactions that contribute to the stability of this large macromolecular machine.
Herpes simplex virus type 1 virions were examined by electron cryomicroscopy, allowing the three-... more Herpes simplex virus type 1 virions were examined by electron cryomicroscopy, allowing the three-dimensional structure of the infectious particle to be visualized for the first time. The capsid shell is identical to that of B-capsids purified from the host cell nucleus, with the exception of the penton channel, which is closed. The double-stranded DNA genome is organized as regularly spaced ( approximately 26 A) concentric layers inside the capsid. This pattern suggests a spool model for DNA packaging, similar to that for some bacteriophages. The bulk of the tegument is not icosahedrally ordered. However, a small portion appears as filamentous structures around the pentons, interacting extensively with the capsid. Their locations and interactions suggest possible roles for the tegument proteins in regulating DNA transport through the penton channel and binding to cellular transport proteins during viral infection.
Electron energy loss spectroscopy and dark-field imaging in a scanning transmission electron micr... more Electron energy loss spectroscopy and dark-field imaging in a scanning transmission electron microscope were used to determine the thickness of glucose-embedded crotoxin complex crystals. The results demonstrate the feasibility of identifying protein crystals with a thickness of half a unit cell (12.8 nm) under low-dose and low-temperature conditions. The accuracy of this method is limited by the amount of surface coating of the crystal's embedding glucose used for preserving the high-resolution structure of the protein. The histogram of the crystal thickness distribution and the spread of the anticipated crystal thickness allow us to make an estimate of the uncertainty in the glucose layer thickness. This approach can be incorporated as part of the experimental procedure in the three-dimensional data collection for structure determination of protein crystals with variable thicknesses. The measurement can be done on areas approximately 200 nm in diameter so that crystals of suit...
Single-particle analysis has become an increasingly important method for structural determination... more Single-particle analysis has become an increasingly important method for structural determination of large macromolecular assemblies. GroEL is an 800 kDa molecular chaperone, which, along with its co-chaperonin GroES, promotes protein folding both in ...
Background: Bluetongue virus (BTV), which belongs to the Reoviridae family and orbivirus genus, i... more Background: Bluetongue virus (BTV), which belongs to the Reoviridae family and orbivirus genus, is a non-enveloped, icosahedral, double-stranded RNA virus. Several protein layers enclose its genome; upon cell entry the outer layer is stripped away leaving a core, the surface of which is composed of VP7. The structure of the trimeric VP7 molecule has previously been determined using X-ray crystallography.
This paper presents a weight function technique for calculating the stress intensity factors for ... more This paper presents a weight function technique for calculating the stress intensity factors for composite repairs to cracks emanating from an internal notch, corrosion blend out, or a free edge under arbitrary loading in rib stiffened panels. The predictions are compared with both finite element and experimental values. This methodology represents a significant extension to existing assessment and design formulae
A three-dimensional reconstruction of a thin protein crystal requires an accurate assignment of a... more A three-dimensional reconstruction of a thin protein crystal requires an accurate assignment of amplitudes and phases in the three-dimensional reciprocal space. This assignment depends upon the tilt parameters which are experimentally determined. A theoretical estimate is made of the tolerable error in the determination of tilt parameters as a function of resolution and crystal thickness.
... ULTRASTRUCTURE RESEARCH 79, 285-293 (1982)Characterization of Crystalline Filtrate Tetanus To... more ... ULTRASTRUCTURE RESEARCH 79, 285-293 (1982)Characterization of Crystalline Filtrate Tetanus ToxinWAH CHIU,*'1 DEBBY RANKERT,* MARY A. CUMMING,t ... We thank Dr. R. Salemme for his discussion.This work has been supported by US Public HealthService Grants ...
Co-chaperonin protein 10 (cpn10, GroES in Escherichia coli) is a ring-shaped heptameric protein t... more Co-chaperonin protein 10 (cpn10, GroES in Escherichia coli) is a ring-shaped heptameric protein that facilitates substrate folding when in complex with cpn60 (GroEL in E. coli). The cpn10 from the hyperthermophilic, ancient bacterium Aquifex aeolicus (Aacpn10) has a 25-residue C-terminal extension in each monomer not found in any other cpn10 protein. Earlier in vitro work has shown that this tail
IN double-stranded-RNA (dsRNA) viruses found in animals1, bacteria2 and yeast3, the genome is tra... more IN double-stranded-RNA (dsRNA) viruses found in animals1, bacteria2 and yeast3, the genome is transcribed within the structurally intact core of the virion with extraordinary efficiency. The structural organization of the genome and the enzymes involved in the transcription inside any of these viruses, critical for understanding this process, is not known. Here we report what we believe is the first
Massive amounts of data are being generated in an effort to represent for the brain the expressio... more Massive amounts of data are being generated in an effort to represent for the brain the expression of all genes at cellular resolution. Critical to exploiting this effort is the ability to place these data into a common frame of reference. Here we have developed a computational method for annotating gene expression patterns in the context of a digital atlas
Proceedings of the National Academy of Sciences of the United States of America, Jan 9, 2015
Lemon-shaped viruses are common in nature but so far have been observed to infect only archaea. D... more Lemon-shaped viruses are common in nature but so far have been observed to infect only archaea. Due to their unusual shape, the structures of these viruses are challenging to study and therefore poorly characterized. Here, we have studied haloarchaeal virus His1 using cryo-electron tomography as well as biochemical dissociation. The virions have different sizes, but prove to be extremely stable under various biochemical treatments. Subtomogram averaging of the computationally extracted virions resolved a tail-like structure with a central tail hub density and six tail spikes. Inside the tail there are two cavities and a plug density that separates the tail hub from the interior genome. His1 most likely uses the tail spikes to anchor to host cells and the tail hub to eject the genome, analogous to classic tailed bacteriophages. Upon biochemical treatment that releases the genome, the lemon-shaped virion transforms into an empty tube. Such a dramatic transformation demonstrates that t...
A program to implement cross-borehole tomography using a simultaneous iterative reconstruction te... more A program to implement cross-borehole tomography using a simultaneous iterative reconstruction technique (SIRT) has been written. The effectiveness of the program in detecting damages in flat plates has been tested through utilising a combination of computer simulations and experimental tests. Images of sample areas were reconstructed from the generation of simulated data for sample areas containing damage zones of different
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