Determination of the accessible surface of globular proteins by means of tritium planigraphy

Results are presented for proteins with known three-dimensional structure (lysozyme, myoglobin, ribonuclease), which show that the probability of label incorporation upon bombardment by hot tritium atoms may be quantitatively linked with the surface area of the protein accessible to water molecules. Possible deviations from simple linear dependency caused by particular mechanisms of label introduction are discussed. The data obtained in experiments with model systems were used to determine the accessible surface area of human serum albumin, for which structural data is not sufficiently accurate to allow estimation of accessible surface area. Experimental data correlate reasonably well with estimations based on conventional concepts of the relationship between accessible surface area and molecular weight for globular proteins. Correspondence to: A. V. Volynskaya     

Structures of Helical Plant Virus Ribonucleoproteins as Assessed by Tritium Planigraphy and Theoretical Modeling

This review considers the results of probing the structure of ribonucleoprotein particles of helical plant viruses by tritium planigraphy (TP). This method works by exposing macromolecular targets to a beam of tritium atoms and analyzing the tritium label distribution along the macromolecule length. The TP data combined with theoretical predictions made it possible to propose a structural model of the coat protein for the virions of potato viruses X (the type representative of potexviruses) and A (a potyvirus), which eluded X-ray diffraction analysis so far. TP revealed fine structural differences between the wild-type tobacco mosaic virus (strain U1) and its temperature-sensitive mutant with an altered coat protein and host specificity. The possibilities of using TP for studying the RNA–protein interactions in helical virus particles are discussed.     

The Spatial Arrangement of the Wild-Type and Mutant Tobacco Mosaic Virus Coat Proteins Assessed by Tritium Planigraphy

Mutant ts21-66 of the tobacco mosaic virus (TMV) differs from the wild-type TMV-U1 by two mutations (Ile21 Thr and Asp66 Gly) in the coat protein (CP) gene and in symptoms produced in infected N" plants. The CP structure in TMV-U1 and ts21-66 virions was probed by tritium planigraphy. Compared with the wild-type CP, labeling of the N-terminal region of mutant CP was half as high and suggested its greater shielding. The role of this CP region in virus interactions with the N" resistance system is discussed.     

Tritium planigraphy: Differences in the spatial structures of the influenza virus M1 protein in crystal,solution, and virion

The structure of the M1 protein of the influenza virus A/Puerto Rico/8/34 (PR8, subtype H1N1) in solution at acidic pH and in the composition of the virion has been studied by the tritium planigraphy method. A model of the spatial structure was constructed using a special algorithm simulating the experiment and a set of algorithms for predicting the secondary structure and disordered regions in proteins. The tertiary structure was refined using the Rosetta program. For a comparison of the structures in solution and inside the virion, the data of X-ray diffraction analysis for the NM domain were also used. The main difference in the structures of the protein in solution and the crystalline state is observed in the region of contact of N and M domains, which in the crystalline state is packed more densely. The regions of the maximum label incorporation almost completely coincide with unstructured regions in the protein that were predicted by the bioinformatics analysis. These regions are concentrated in the C domain and in loop regions between M, N, and C domains. The data were confirmed by analytical centrifugation and dynamic light scattering. Anomalous hydrodynamic dimensions and a low structuration of the M1 protein in solution were found. The polyfunctionality of the protein in the cell is probably related to its flexible tertiary structure, which, owing to unstructured regions, provides contact with various partner molecules.     

Spectral fluorescence: an ataxonomic tool for studying the structure of phytoplankton populations

Although early studies in plankton biology feature detailedanalyses of numbers of species, modern studies tend to emphasizebulk changes in biomass associated with rare processes. We arguethat the penalty for this approach will eventually be an incompleteunderstanding of seasonal and spatial changes in the patternof primary production of ocean waters. The ability of naturalpopulations to respond to changes in the marine environmentis due to adaptability at the cellular level (phenotypic) andchanges in whole species groups (genotypic). The inability totest the relative importance of these two strategies is dueto the incompatiblity of traditional taxonomic approaches andfield experimentation where rate processes are being measured.To counteract this difficulty we propose an ataxonomic techniquewhich utilizes the differences in light absorption by phytoplanktonfor photosynthetic processes. In this report we review how changesin the fluorescence spectral signatures reflect other biochemicalfeatures in natural populations. This is examined in concertwith physical and chemical changes associated with the characteristicsof ocean water masses with specific reference as to how thesehydrographic features influence the relative growth of phytoplanktonpopulations. ^*This paper is the result of a study made at the Group for AquaticPrimaiy Productivity (GAP). Second International Workshop heldat the National Oceanographic Institute, Haifa, Israel in April–May1984.     

DSSR: an integrated software tool for dissecting the spatial structure of RNA

Insight into the three-dimensional architecture of RNA is essential for understanding its cellular functions. However, even the classic transfer RNA structure contains features that are overlooked by existing bioinformatics tools. Here we present DSSR (Dissecting the Spatial Structure of RNA), an integrated and automated tool for analyzing and annotating RNA tertiary structures. The software identifies canonical and noncanonical base pairs, including those with modified nucleotides, in any tautomeric or protonation state. DSSR detects higher-order coplanar base associations, termed multiplets. It finds arrays of stacked pairs, classifies them by base-pair identity and backbone connectivity, and distinguishes a stem of covalently connected canonical pairs from a helix of stacked pairs of arbitrary type/linkage. DSSR identifies coaxial stacking of multiple stems within a single helix and lists isolated canonical pairs that lie outside of a stem. The program characterizes ‘closed’ loops of various types (hairpin, bulge, internal, and junction loops) and pseudoknots of arbitrary complexity. Notably, DSSR employs isolated pairs and the ends of stems, whether pseudoknotted or not, to define junction loops. This new, inclusive definition provides a novel perspective on the spatial organization of RNA. Tests on all nucleic acid structures in the Protein Data Bank confirm the efficiency and robustness of the software, and applications to representative RNA molecules illustrate its unique features. DSSR and related materials are freely available at http://x3dna.org/.     

Study of the spatial structure of globular proteins by tritium planigraphy. Short peptides as a model of a fully extended polypeptide chain]

The interaction of tritium atoms with amino acid residue from short peptides was studied. The short peptides were considered as a model of extended polypeptides chain. Every residue in this chain has 100% steric accessibility. It was shown that: 1. The linear correlation exists between the residue accessible surface area (that is composed of hydrocarbon fragments) and the amount of tritium interacting with this residue; 2. The presence of the tertiary carbon atom in the residue side chain influences on the reactivity of this residue; 3. The N- or C-terminal residue presence does not influences on the possibility of interaction of this residue with tritium atoms. The obtained reactivity scale of amino acid residues is compared with other theoretical and experimental data.     

GLASS: A tool to visualize protein structure prediction data in three dimensions and evaluate their consistency

When a protein sequence does not share any significant sequence similarity with a protein of known structure, homology modeling cannot be applied. However, many novel and interesting methods, such as secondary structure prediction, fold recognition, and prediction of long-range interactions, are being developed and have been shown to be reasonably successful in predicting protein structures from sequence data and evolutionary information. The a priori evaluation of the correctness of a prediction obtained by one of these methods is however often problematic. Consequently, it is important to use all available information provided by as many different methods as possible and all the available experimental data about the protein of interest, since the consistency of the results is indicative of the reliability of the prediction. Hence the need has arisen for suitable tools able to compare results provided by different methods and evaluate their consistency. We have therefore constructed GLASS, a general platform to read, visualize, compare, and evaluate prediction results from many different sources and to project these prediction results into three dimensions. In addition, GLASS allows the comparison of selected parameters calculated for a model with the distribution observed in real protein structures, thus providing an easy way to test new methods for evaluating the likelihood of different structural models. GLASS can be considered as a “workbench” for structural predictions useful to both experimentalists and theoreticians. Proteins 30:339–351, 1998. © 1998 Wiley-Liss, Inc.     

Magic angle spinning (MAS) NMR: a new tool to study the spatial and electronic structure of photosynthetic complexes

Photosynthesis Research - In the last two decades, Magic Angle Spinning (MAS) NMR has created its own niche in studies involving photosynthetic membrane protein complexes, owing to its ability to...     

A microsatellite DNA toolkit for studying population structure in Apis mellifera

We present a set of 18 microsatellite DNA markers that can be run in two multiplex polymerase chain reactions as standard tool for assessing molecular ecological problems in honeybees (Apis mellifera). In addition to a set of six unlinked loci testing for classical population genetic parameters, we present three sets of four tightly linked loci, each located on three different chromosomes. These linked markers are useful for determining the number of colonies in a population as well as the parentage of drones and workers. Moreover, the tool kit can test for various modes of natural selection in honeybee populations.     

A protein short motif search tool using amino acid sequence and their secondary structure assignment

We present the development of a web server, a protein short motif search tool that allows users to simultaneously search for a protein sequence motif and its secondary structure assignments. The web server is able to query very short motifs searches against PDB structural data from the RCSB Protein Databank, with the users defining the type of secondary structures of the amino acids in the sequence motif. The output utilises 3D visualisation ability that highlights the position of the motif in the structure and on the corresponding sequence. Researchers can easily observe the locations and conformation of multiple motifs among the results. Protein short motif search also has an application programming interface (API) for interfacing with other bioinformatics tools. AVAILABILITY: The database is available for free at http://birg3.fbb.utm.my/proteinsms.     

Comparative sequence analysis as a tool for studying the secondary structure of mRNAs.

Analysis of phylogenetically conserved secondary structure has been important in the development of models for the secondary structure of structural RNAs. In this paper, we apply this type of analysis to several families of informational RNAs to evaluate its usefulness in developing secondary structure models for mRNAs and mRNA precursors. We observed many conserved helices in all mRNA groups analyzed. Three criteria were used to identify potential helices which were not conserved solely because of coding sequence constraints, and may therefore be important for the structure and function of the RNA. These results suggest that this approach will be useful in deriving secondary structure models for informational RNAs when used in conjunction with other complementary techniques, and in designing experiments to determine the functional significance of conserved base pairing interactions.     

Protein-nucleic acid complexes and the role of mass spectrometry in their structure determination

Mass spectrometry is now established as a powerful tool for the study of the stoichiometry, interactions, dynamics, and subunit architecture of large protein assemblies and their subcomplexes. Recent evidence has suggested that the 3D structure of protein complexes can be maintained intact in the gas phase, highlighting the potential of ion mobility to contribute to structural biology. A key challenge is to integrate the compositional and structural information from ion mobility mass spectrometry with molecular modelling approaches to produce 3D models of intact protein complexes. In this review, we focus on the mass spectrometry of protein-nucleic acid assemblies with particular attention to the application of ion mobility, an emerging technique in structural studies. We also discuss the challenges that lie ahead for the full integration of ion mobility mass spectrometry with structural biology.     

Single particle reconstruction of membrane proteins: A tool for understanding the 3D structure of disease-related macromolecules

Membrane proteins play important roles in cell functions such as neurotransmission, muscle contraction, and hormone secretion, but their structures are mostly undetermined. Several techniques have been developed to elucidate the structure of macromolecules; X-ray or electron crystallography, nuclear magnetic resonance spectroscopy, and high-resolution electron microscopy. Electron microscopy-based single particle reconstruction, a computer-aided structure determination method, reconstructs a three-dimensional (3D) structure from projections of monodispersed protein. A large number of particle images are picked up from EM films, aligned and classified to generate two-dimensional (2D) averages, and, using the Euler angle of each 2D average, reconstructed into a 3D structure. This method is challenging due to the necessity for close collaboration between classical biochemistry and innovative information technology, including parallel computing. However, recent progress in electron microscopy, mathematical algorithms, and computational ability has greatly increased the subjects that are considered to be primarily addressable using single particle reconstruction. Membrane proteins are one of these targets to which the single particle reconstruction is successfully applied for understanding of their structures. In this paper, we will introduce recently reconstructed channel-related proteins and discuss the applicability of this technique in understanding molecular structures and their roles in pathology.     

The SPASIBA force field as an essential tool for studying the structure and dynamics of saccharides

The SPASIBA force field has been applied to the determination of the structure and dynamical properties of various disaccharides. It has been shown that the experimental properties (structure, dipole moment, conformational relative energies) are satisfactorily predicted. The anomeric and exo-anomeric effects are confidently reproduced without specific terms for the alpha and beta anomers and the type of glycosidic linkages.     

Tritium labelling of the amino acids of Sinapis alba is identical in darkness and far-red light

Earlier reports that irradiation for 3 hr with continuous far-red light stimulated the incorporation of deuterium from deuterium oxide into the free amino acids of the cotyledons of Sinapis alba have not been confirmed by studies using tritium oxide. Possible explanations of the discrepancy are discussed.     

TCP: a tool for designing chimera proteins based on the tertiary structure information

Background  

Chimera proteins are widely used for the analysis of the protein-protein interaction region. One of the major issues is the epitope analysis of the monoclonal antibody. In the analysis, a continuous portion of an antigen is sequentially substituted into a different sequence. This method works well for an antibody recognizing a linear epitope, but not for that recognizing a discontinuous epitope. Although the designing the chimera proteins based on the tertiary structure information is required in such situations, there is no appropriate tool so far.