Proteomic analysis of the bacterial cell cycle

A global approach was used to analyze protein synthesis and stability during the cell cycle of the bacterium Caulobacter crescentus. Approximately one-fourth (979) of the estimated C. crescentus gene products were detected by two-dimensional gel electrophoresis, 144 of which showed differential cell cycle expression patterns. Eighty-one of these proteins were identified by mass spectrometry and were assigned to a wide variety of functional groups. Pattern analysis revealed that coexpression groups were functionally clustered. A total of 48 proteins were rapidly degraded in the course of one cell cycle. More than half of these unstable proteins were also found to be synthesized in a cell cycle-dependent manner, establishing a strong correlation between rapid protein turnover and the periodicity of the bacterial cell cycle. This is, to our knowledge, the first evidence for a global role of proteolysis in bacterial cell cycle control.

Molecular Cloning and Expression Analysis of the Mitochondrial Pyruvate Dehydrogenase from Maize1

Four cDNAs, one encoding an α-subunit and three encoding β-subunits of the mitochondrial pyruvate dehydrogenase, were isolated from maize (Zea mays L.) libraries. The deduced amino acid sequences of both α- and β-subunits are approximately 80% identical with Arabidopsis and pea (Pisum sativum L.) homologs. The mature N terminus was determined for the β-subunit by microsequencing the protein purified from etiolated maize shoot mitochondria and was resolved by two-dimensional gel electrophoresis. This single isoelectric species comprised multiple isoforms. Both α- and β-subunits are encoded by multigene families in maize, as determined by Southern-blot analyses. RNA transcripts for both α- and β-subunits were more abundant in roots than in young leaves or etiolated shoots. Pyruvate dehydrogenase activity was also higher in roots (5-fold) compared with etiolated shoots and leaves. Both subunits were present at similar levels in all tissues examined, indicating coordinated gene regulation. The protein levels were highest in heterotrophic organs and in pollen, which contained about 2-fold more protein than any other organ examined. The relative abundance of these proteins in nonphotosynthetic tissues may reflect a high cellular content of mitochondria, a high level of respiratory activity, or an extra plastidial requirement for acetate.

Photoactive yellow protein: A structural prototype for the three-dimensional fold of the PAS domain superfamily

PAS domains are found in diverse proteins throughout all three kingdoms of life, where they apparently function in sensing and signal transduction. Although a wealth of useful sequence and functional information has become recently available, these data have not been integrated into a three-dimensional (3D) framework. The very early evolutionary development and diverse functions of PAS domains have made sequence analysis and modeling of this protein superfamily challenging. Limited sequence similarities between the ∼50-residue PAS repeats and one region of the bacterial blue-light photosensor photoactive yellow protein (PYP), for which ground-state and light-activated crystallographic structures have been determined to high resolution, originally were identified in sequence searches using consensus sequence probes from PAS-containing proteins. Here, we found that by changing a few residues particular to PYP function, the modified PYP sequence probe also could select PAS protein sequences. By mapping a typical ∼150-residue PAS domain sequence onto the entire crystallographic structure of PYP, we show that the PAS sequence similarities and differences are consistent with a shared 3D fold (the PAS/PYP module) with obvious potential for a ligand-binding cavity. Thus, PYP appears to prototypically exhibit all the major structural and functional features characteristic of the PAS domain superfamily: the shared PAS/PYP modular domain fold of ∼125–150 residues, a sensor function often linked to ligand or cofactor (chromophore) binding, and signal transduction capability governed by heterodimeric assembly (to the downstream partner of PYP). This 3D PAS/PYP module provides a structural model to guide experimental testing of hypotheses regarding ligand-binding, dimerization, and signal transduction.

Statistical analysis of shape of objects based on landmark data.

Two objects with homologous landmarks are said to be of the same shape if the configurations of landmarks of one object can be exactly matched with that of the other by translation, rotation/reflection, and scaling. The observations on an object are coordinates of its landmarks with reference to a set of orthogonal coordinate axes in an appropriate dimensional space. The origin, choice of units, and orientation of the coordinate axes with respect to an object may be different from object to object. In such a case, how do we quantify the shape of an object, find the mean and variation of shape in a population of objects, compare the mean shapes in two or more different populations, and discriminate between objects belonging to two or more different shape distributions. We develop some methods that are invariant to translation, rotation, and scaling of the observations on each object and thereby provide generalizations of multivariate methods for shape analysis.

Characterizations of diverse residue clusters in protein three-dimensional structures.

We present new methods for identifying and analyzing statistically significant residue clusters that occur in three-dimensional (3D) protein structures. Residue clusters of different kinds occur in many contexts. They often feature the active site (e.g., in substrate binding), the interface between polypeptide units of protein complexes, regions of protein-protein and protein-nucleic acid interactions, or regions of metal ion coordination. The methods are illustrated with 3D clusters centering on four themes. (i) Acidic or histidine-acidic clusters associated with metal ions. (ii) Cysteine clusters including coordination of metals such as zinc or iron-sulfur structures, cysteine knots prominent in growth factors, multiple sets of buried disulfide pairings that putatively nucleate the hydrophobic core, or cysteine clusters of mostly exposed disulfide bridges. (iii) Iron-sulfur proteins and charge clusters. (iv) 3D environments of multiple histidine residues. Study of diverse 3D residue clusters offers a new perspective on protein structure and function. The algorithms can aid in rapid identification of distinctive sites, suggest correlations among protein structures, and serve as a tool in the analysis of new structures.

Two-dimensional protein crystallization via metal-ion coordination by naturally occurring surface histidines.

A powerful and potentially general approach to the targeting and crystallization of proteins on lipid interfaces through coordination of surface histidine residues to lipid-chelated divalent metal ions is presented. This approach, which should be applicable to the crystallization of a wide range of naturally occurring or engineered proteins, is illustrated here by the crystallization of streptavidin on a monolayer of an iminodiacetate-Cu(II) lipid spread at the air-water interface. This method allows control of the protein orientation at interfaces, which is significant for the facile production of highly ordered protein arrays and for electron density mapping in structural analysis of two-dimensional crystals. Binding of native streptavidin to the iminodiacetate-Cu lipids occurs via His-87, located on the protein surface near the biotin binding pocket. The two-dimensional streptavidin crystals show a previously undescribed microscopic shape that differs from that of crystals formed beneath biotinylated lipids.

Observation via one-dimensional 13Calpha NMR of local conformational substates in thermal unfolding equilibria of a synthetic analog of the GCN4 leucine zipper.

Synthesis of a 33-residue, capped leucine zipper analogous to that in GCN4 is reported. Histidine and arginine residues are mutated to lysine to reduce the unfolding temperature. CD and ultracentrifugation studies indicate that the molecule is a two-stranded coiled coil under benign conditions. Versions of the same peptide are made with 99% 13Calpha at selected sites. One-dimensional 13C NMR spectra are assigned by inspection and used to study thermal unfolding equilibria over the entire transition from 8 to 73 degrees C. Spectra at the temperature extremes establish the approximate chemical shifts for folded and unfolded forms at each labeled site. Resonances for each amino acid appear at both locations at intermediate T, indicating that folded and unfolded forms interconvert slowly (> >2 ms) on the NMR time scale. Moreover, near room temperature, the structured form's resonance is double at several, but not all, sites, indicating at least two slowly interconverting, structured, local conformational substates. Analysis of the dynamics is possible. For example, near room temperature at the Val-9, Ala-24, and Gly-31 positions, the equilibrium constant for interconversion of the two structured forms is near unity and the time scale is > or= 10-20 ms.

Finite-size scaling analysis of the distributions of pseudo-critical temperatures in spin glasses

Using the results of large scale numerical simulations we study the probability distribution of the pseudo critical temperature for the three dimensional Edwards Anderson Ising spin glass and for the fully connected Sherrington-Kirkpatrick model. We find that the behaviour of our data is nicely described by straightforward finitesize scaling relations.

Human Motion Analysis Based on Sequential Modeling of Radar Signal and Stereo Image Features

Falls are one of the greatest threats to elderly health in their daily living routines and activities. Therefore, it is very important to detect falls of an elderly in a timely and accurate manner, so that immediate response and proper care can be provided, by sending fall alarms to caregivers. Radar is an effective non-intrusive sensing modality which is well suited for this purpose, which can detect human motions in all types of environments, penetrate walls and fabrics, preserve privacy, and is insensitive to lighting conditions. Micro-Doppler features are utilized in radar signal corresponding to human body motions and gait to detect falls using a narrowband pulse-Doppler radar. Human motions cause time-varying Doppler signatures, which are analyzed using time-frequency representations and matching pursuit decomposition (MPD) for feature extraction and fall detection. The extracted features include MPD features and the principal components of the time-frequency signal representations. To analyze the sequential characteristics of typical falls, the extracted features are used for training and testing hidden Markov models (HMM) in different falling scenarios. Experimental results demonstrate that the proposed algorithm and method achieve fast and accurate fall detections. The risk of falls increases sharply when the elderly or patients try to exit beds. Thus, if a bed exit can be detected at an early stage of this motion, the related injuries can be prevented with a high probability. To detect bed exit for fall prevention, the trajectory of head movements is used for recognize such human motion. A head detector is trained using the histogram of oriented gradient (HOG) features of the head and shoulder areas from recorded bed exit images. A data association algorithm is applied on the head detection results to eliminate head detection false alarms. Then the three dimensional (3D) head trajectories are constructed by matching scale-invariant feature transform (SIFT) keypoints in the detected head areas from both the left and right stereo images. The extracted 3D head trajectories are used for training and testing an HMM based classifier for recognizing bed exit activities. The results of the classifier are presented and discussed in the thesis, which demonstrates the effectiveness of the proposed stereo vision based bed exit detection approach.

Analysis of the colorimetric properties of goniochromatic colors using the MacAdam limits under different light sources

Technological innovation in all areas has led to the appearance in recent years of new metallic and pearlescent materials, yet no exhaustive studies have been conducted to assess their colorimetric capabilities. The chromatic variability of these special-effect pigments may largely be due to the three-dimensional effect of their curved shapes and orientations when they are directionally or diffusely illuminated. Our study examines goniochromatic colors using the optimal colors (MacAdam limits) associated with normal colors (photometric scale of relative spectral reflectance from 0 to 1) under certain conventional illuminants and other light sources. From a database of 91 metallic and interference samples and using a multi-gonio-spectrophotometer, we analyzed samples with lightness values of more than 100 and others with lightness values of less than 100, but with higher chromaticities than optimal colors, which places them beyond the MacAdam limits. Our study thus demonstrates the existence of chromatic perceptions beyond the normal solid color associated with these materials and independent of the light source. The challenge for future research, therefore, is to replicate and render these color appearances in current and future color reproduction technologies for computer graphics.

Spin degree of freedom in two dimensional exciton condensates

We present a theoretical analysis of a spin-dependent multicomponent condensate in two dimensions. The case of a condensate of resonantly photoexcited excitons having two different spin orientations is studied in detail. The energy and the chemical potentials of this system depend strongly on the spin polarization. When electrons and holes are located in two different planes, the condensate can be either totally spin polarized or spin unpolarized, a property that is measurable. The phase diagram in terms of the total density and electron-hole separation is discussed.

Discontinuity spacing analysis in rock masses using 3D point clouds

The complete characterization of rock masses implies the acquisition of information of both, the materials which compose the rock mass and the discontinuities which divide the outcrop. Recent advances in the use of remote sensing techniques – such as Light Detection and Ranging (LiDAR) – allow the accurate and dense acquisition of 3D information that can be used for the characterization of discontinuities. This work presents a novel methodology which allows the calculation of the normal spacing of persistent and non-persistent discontinuity sets using 3D point cloud datasets considering the three dimensional relationships between clusters. This approach requires that the 3D dataset has been previously classified. This implies that discontinuity sets are previously extracted, every single point is labeled with its corresponding discontinuity set and every exposed planar surface is analytically calculated. Then, for each discontinuity set the method calculates the normal spacing between an exposed plane and its nearest one considering 3D space relationship. This link between planes is obtained calculating for every point its nearest point member of the same discontinuity set, which provides its nearest plane. This allows calculating the normal spacing for every plane. Finally, the normal spacing is calculated as the mean value of all the normal spacings for each discontinuity set. The methodology is validated through three cases of study using synthetic data and 3D laser scanning datasets. The first case illustrates the fundamentals and the performance of the proposed methodology. The second and the third cases of study correspond to two rock slopes for which datasets were acquired using a 3D laser scanner. The second case study has shown that results obtained from the traditional and the proposed approaches are reasonably similar. Nevertheless, a discrepancy between both approaches has been found when the exposed planes members of a discontinuity set were hard to identify and when the planes pairing was difficult to establish during the fieldwork campaign. The third case study also has evidenced that when the number of identified exposed planes is high, the calculated normal spacing using the proposed approach is minor than those using the traditional approach.

Three-dimensional planar object tracking with sub-pixel accuracy

Subpixel techniques are commonly used to increase the spatial resolution in tracking tasks. Object tracking with targets of known shape permits obtaining information about object position and orientation in the three-dimensional space. A proper selection of the target shape allows us to determine its position inside a plane and its angular and azimuthal orientation under certain limits. Our proposal is demonstrated both numerical and experimentally and provides an increase the accuracy of more than one order of magnitude compared to the nominal resolution of the sensor. The experiment has been performed with a high-speed camera, which simultaneously provides high spatial and temporal resolution, so it may be interesting for some applications where this kind of targets can be attached, such as vibration monitoring and structural analysis.

Mapping Emotion Terms into Affective Space: Further Evidence for a Four-Dimensional Structure

Recent empirical work on the semantics of emotion terms across many different cultures and languages, using a theoretical componential approach, suggested that four dimensions are needed to parsimoniously describe the semantic space of the emotion domain as reflected in emotion terms (Fontaine, Scherer, Roesch, & Ellsworth, 2007; Fontaine, Scherer, & Soriano, 2013). In addition to valence, power, and arousal, a novelty dimension was discovered that mostly differentiated surprise from other emotions. Here, we further explore the existence and nature of the fourth dimension in semantic emotion space using a much larger and much more representative set of emotion terms. A group of 156 participants each rated 10 out of a set of 80 French emotion terms with respect to semantic meaning. The meaning of an emotion term was evaluated with respect to 68 emotion features representing the appraisal, action tendency, bodily reaction, expression, and feeling components of the emotion process. A principal component analysis confirmed the four-dimensional valence, power, arousal, and novelty structure. Moreover, this larger and much more representative set of emotion terms revealed that the novelty dimension not only differentiates surprise terms from other emotion terms, but also identifies substantial variation within the fear and joy emotion families.

Spectral analysis of seabed elevation from the Danish Wadden Sea

Bedforms both reflect and influence shallow water hydrodynamics and sediment dynamics. A correct characterization of their spatial distribution and dimensions is required for the understanding, assessment and prediction of numerous coastal processes. A method to parameterize geometrical characteristics using two-dimensional (2D) spectral analysis is presented and tested on seabed elevation data from the Knudedyb tidal inlet in the Danish Wadden Sea, where large compound bedforms are found. The bathymetric data were divided into 20x20 m areas on which a 2D spectral analysis was applied. The most energetic peak of the 2D spectrum was found and its energy, frequency and direction were calculated. A power-law was fitted to the average of slices taken through the 2D spectrum; its slope and y-intercept were calculated. Using these results the test area was morphologically classified into 4 distinct morphological regions. The most energetic peak and the slope and intercept of the power-law showed high values above the crest of the primary bedforms and scour holes, low values in areas without bedforms, and intermediate values in areas with secondary bedforms. The secondary bedform dimensions and orientations were calculated. An area of 700x700 m was used to determine the characteristics of the primary bedforms. However, they were less distinctively characterized compared to the secondary bedforms due to relatively large variations in their orientations and wavelengths. The method is thus appropriate for morphological classification of the seabed and for bedform characterization, being most efficient in areas characterized by bedforms with regular dimensions and directions.