Large-scale protein structure modeling of the Saccharomyces cerevisiae genome

The function of a protein generally is determined by its three-dimensional (3D) structure. Thus, it would be useful to know the 3D structure of the thousands of protein sequences that are emerging from the many genome projects. To this end, fold assignment, comparative protein structure modeling, and model evaluation were automated completely. As an illustration, the method was applied to the proteins in the Saccharomyces cerevisiae (baker’s yeast) genome. It resulted in all-atom 3D models for substantial segments of 1,071 (17%) of the yeast proteins, only 40 of which have had their 3D structure determined experimentally. Of the 1,071 modeled yeast proteins, 236 were related clearly to a protein of known structure for the first time; 41 of these previously have not been characterized at all.

Three-dimensional modeling of and ligand docking to vitamin D receptor ligand binding domain

The ligand binding domain of the human vitamin D receptor (VDR) was modeled based on the crystal structure of the retinoic acid receptor. The ligand binding pocket of our VDR model is spacious at the helix 11 site and confined at the β-turn site. The ligand 1α,25-dihydroxyvitamin D3 was assumed to be anchored in the ligand binding pocket with its side chain heading to helix 11 (site 2) and the A-ring toward the β-turn (site 1). Three residues forming hydrogen bonds with the functionally important 1α- and 25-hydroxyl groups of 1α,25-dihydroxyvitamin D3 were identified and confirmed by mutational analysis: the 1α-hydroxyl group is forming pincer-type hydrogen bonds with S237 and R274 and the 25-hydroxyl group is interacting with H397. Docking potential for various ligands to the VDR model was examined, and the results are in good agreement with our previous three-dimensional structure-function theory.

Molecular phylogenetic analysis of evolutionary trends in stonefly wing structure and locomotor behavior

Insects in the order Plecoptera (stoneflies) use a form of two-dimensional aerodynamic locomotion called surface skimming to move across water surfaces. Because their weight is supported by water, skimmers can achieve effective aerodynamic locomotion even with small wings and weak flight muscles. These mechanical features stimulated the hypothesis that surface skimming may have been an intermediate stage in the evolution of insect flight, which has perhaps been retained in certain modern stoneflies. Here we present a phylogeny of Plecoptera based on nucleotide sequence data from the small subunit rRNA (18S) gene. By mapping locomotor behavior and wing structural data onto the phylogeny, we distinguish between the competing hypotheses that skimming is a retained ancestral trait or, alternatively, a relatively recent loss of flight. Our results show that basal stoneflies are surface skimmers, and that various forms of surface skimming are distributed widely across the plecopteran phylogeny. Stonefly wings show evolutionary trends in the number of cross veins and the thickness of the cuticle of the longitudinal veins that are consistent with elaboration and diversification of flight-related traits. These data support the hypothesis that the first stoneflies were surface skimmers, and that wing structures important for aerial flight have become elaborated and more diverse during the radiation of modern stoneflies.

Analysis of six prophages in Lactococcus lactis IL1403: different genetic structure of temperate and virulent phage populations

We report the genetic organisation of six prophages present in the genome of Lactococcus lactis IL1403. The three larger prophages (36–42 kb), belong to the already described P335 group of temperate phages, whereas the three smaller ones (13–15 kb) are most probably satellites relying on helper phage(s) for multiplication. These data give a new insight into the genetic structure of lactococcal phage populations. P335 temperate phages have variable genomes, sharing homology over only 10–33% of their length. In contrast, virulent phages have highly similar genomes sharing homology over >90% of their length. Further analysis of genetic structure in all known groups of phages active on other bacterial hosts such as Escherichia coli, Bacillus subtilis, Mycobacterium and Streptococcus thermophilus confirmed the existence of two types of genetic structure related to the phage way of life. This might reflect different intensities of horizontal DNA exchange: low among purely virulent phages and high among temperate phages and their lytic homologues. We suggest that the constraints on genetic exchange among purely virulent phages reflect their optimal genetic organisation, adapted to a more specialised and extreme form of parasitism than temperate/lytic phages.

X-ray structure of the human hyperplastic discs protein: An ortholog of the C-terminal domain of poly(A)-binding protein

The poly(A)-binding protein (PABP) recognizes the 3′ mRNA poly(A) tail and plays an essential role in eukaryotic translation initiation and mRNA stabilization/degradation. PABP is a modular protein, with four N-terminal RNA-binding domains and an extensive C terminus. The C-terminal region of PABP is essential for normal growth in yeast and has been implicated in mediating PABP homo-oligomerization and protein–protein interactions. A small, proteolytically stable, highly conserved domain has been identified within this C-terminal segment. Remarkably, this domain is also present in the hyperplastic discs protein (HYD) family of ubiquitin ligases. To better understand the function of this conserved region, an x-ray structure of the PABP-like segment of the human HYD protein has been determined at 1.04-Å resolution. The conserved domain adopts a novel fold resembling a right-handed supercoil of four α-helices. Sequence profile searches and comparative protein structure modeling identified a small ORF from the Arabidopsis thaliana genome that encodes a structurally similar but distantly related PABP/HYD domain. Phylogenetic analysis of the experimentally determined (HYD) and homology modeled (PABP) protein surfaces revealed a conserved feature that may be responsible for binding to a PABP interacting protein, Paip1, and other shared interaction partners.

Mutational analysis and molecular modeling of the nonapeptide hormone binding domains of the [Arg8]vasotocin receptor.

To identify determinants that form nonapeptide hormone binding domains of the white sucker Catostomus commersoni [Arg8]vasotocin receptor, chimeric constructs encoding parts of the vasotocin receptor and parts of the isotocin receptor have been analyzed by [(3,5-3H)Tyr2, Arg8]vasotocin binding to membranes of human embryonic kidney cells previously transfected with the different cDNA constructs and by functional expression studies in Xenopus laevis oocytes injected with mutant cRNAs. The results indicate that the N terminus and a region spanning the second extracellular loop and its flanking transmembrane segments, which contains a number of amino acid residues that are conserved throughout the nonapeptide receptor family, contribute to the affinity of the receptor for its ligand. Nonapeptide selectivity, however, is mainly defined by transmembrane region VI and the third extracellular loop. These results are complemented by a molecular model of the vasotocin receptor obtained by aligning its sequence with those of other G-protein coupled receptors as well as that of bacteriorhodopsin. The model indicates that amino acid residues of transmembrane regions II-VII that are located close to the extracellular surface also contribute to the binding of vasotocin.

Structure-activity analysis of thanatin, a 21-residue inducible insect defense peptide with sequence homology to frog skin antimicrobial peptides.

Immune challenge to the insect Podisus maculiventris induces synthesis of a 21-residue peptide with sequence homology to frog skin antimicrobial peptides of the brevinin family. The insect and frog peptides have in common a C-terminally located disulfide bridge delineating a cationic loop. The peptide is bactericidal and fungicidal, exhibiting the largest antimicrobial spectrum observed so far for an insect defense peptide. An all-D-enantiomer is nearly inactive against Gram-negative bacteria and some Gram-positive strains but is fully active against fungi and other Gram-positive bacteria, suggesting that more than one mechanism accounts for the antimicrobial activity of this peptide. Studies with truncated synthetic isoforms underline the role of the C-terminal loop and flanking residues for the activity of this molecule for which we propose the name thanatin.

Analysis of homeodomain function by structure-based design of a transcription factor.

The homeodomain is a 60-amino acid module which mediates critical protein-DNA and protein-protein interactions for a large family of regulatory proteins. We have used structure-based design to analyze the ability of the Oct-1 homeodomain to nucleate an enhancer complex. The Oct-1 protein regulates herpes simplex virus (HSV) gene expression by participating in the formation of a multiprotein complex (C1 complex) which regulates alpha (immediate early) genes. We recently described the design of ZFHD1, a chimeric transcription factor containing zinc fingers 1 and 2 of Zif268, a four-residue linker, and the Oct-1 homeodomain. In the presence of alpha-transinduction factor and C1 factor, ZFHD1 efficiently nucleates formation of the C1 complex in vitro and specifically activates gene expression in vivo. The sequence specificity of ZFHD1 recruits C1 complex formation to an enhancer element which is not efficiently recognized by Oct-1. ZFHD1 function depends on the recognition of the Oct-1 homeodomain surface. These results prove that the Oct-1 homeodomain mediates all the protein-protein interactions that are required to efficiently recruit alpha-transinduction factor and C1 factor into a C1 complex. The structure-based design of transcription factors should provide valuable tools for dissecting the interactions of DNA-bound domains in other regulatory circuits.

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.

Dynamic investigation of an ancient masonry bell tower with operational modal analysis: a non-destructive experimental technique to obtain the dynamic characteristics of a structure

This paper shows the results of an experimental analysis on the bell tower of “Chiesa della Maddalena” (Mola di Bari, Italy), to better understand the structural behavior of slender masonry structures. The research aims to calibrate a numerical model by means of the Operational Modal Analysis (OMA) method. In this way realistic conclusions about the dynamic behavior of the structure are obtained. The choice of using an OMA derives from the necessity to know the modal parameters of a structure with a non-destructive testing, especially in case of cultural-historical value structures. Therefore by means of an easy and accurate process, it is possible to acquire in-situ environmental vibrations. The data collected are very important to estimate the mode shapes, the natural frequencies and the damping ratios of the structure. To analyze the data obtained from the monitoring, the Peak Picking method has been applied to the Fast Fourier Transforms (FFT) of the signals in order to identify the values of the effective natural frequencies and damping factors of the structure. The main frequencies and the damping ratios have been determined from measurements at some relevant locations. The responses have been then extrapolated and extended to the entire tower through a 3-D Finite Element Model. In this way, knowing the modes of vibration, it has been possible to understand the overall dynamic behavior of the structure.

Logical Modeling and Dynamical Analysis of Cellular Networks

The logical (or logic) formalism is increasingly used to model regulatory and signaling networks. Complementing these applications, several groups contributed various methods and tools to support the definition and analysis of logical models. After an introduction to the logical modeling framework and to several of its variants, we review here a number of recent methodological advances to ease the analysis of large and intricate networks. In particular, we survey approaches to determine model attractors and their reachability properties, to assess the dynamical impact of variations of external signals, and to consistently reduce large models. To illustrate these developments, we further consider several published logical models for two important biological processes, namely the differentiation of T helper cells and the control of mammalian cell cycle.