Mechanical Strength of 17 134 Model Proteins and Cysteine Slipknots

A new theoretical survey of proteins' resistance to constant speed stretching is performed for a set of 17 134 proteins as described by a structure-based model. The proteins selected have no gaps in their structure determination and consist of no more than 250 amino acids. Our previous studies have dealt with 7510 proteins of no more than 150 amino acids. The proteins are ranked according to the strength of the resistance. Most of the predicted top-strength proteins have not yet been studied experimentally. Architectures and folds which are likely to yield large forces are identified. New types of potent force clamps are discovered. They involve disulphide bridges and, in particular, cysteine slipknots. An effective energy parameter of the model is estimated by comparing the theoretical data on characteristic forces to the corresponding experimental values combined with an extrapolation of the theoretical data to the experimental pulling speeds. These studies provide guidance for future experiments on single molecule manipulation and should lead to selection of proteins for applications. A new class of proteins, involving cystein slipknots, is identified as one that is expected to lead to the strongest force clamps known. This class is characterized through molecular dynamics simulations.Comment: 40 pages, 13 PostScript figure

Two Pfam protein families characterized by a crystal structure of protein lpg2210 from Legionella pneumophila.

BackgroundEvery genome contains a large number of uncharacterized proteins that may encode entirely novel biological systems. Many of these uncharacterized proteins fall into related sequence families. By applying sequence and structural analysis we hope to provide insight into novel biology.ResultsWe analyze a previously uncharacterized Pfam protein family called DUF4424 [Pfam:PF14415]. The recently solved three-dimensional structure of the protein lpg2210 from Legionella pneumophila provides the first structural information pertaining to this family. This protein additionally includes the first representative structure of another Pfam family called the YARHG domain [Pfam:PF13308]. The Pfam family DUF4424 adopts a 19-stranded beta-sandwich fold that shows similarity to the N-terminal domain of leukotriene A-4 hydrolase. The YARHG domain forms an all-helical domain at the C-terminus. Structure analysis allows us to recognize distant similarities between the DUF4424 domain and individual domains of M1 aminopeptidases and tricorn proteases, which form massive proteasome-like capsids in both archaea and bacteria.ConclusionsBased on our analyses we hypothesize that the DUF4424 domain may have a role in forming large, multi-component enzyme complexes. We suggest that the YARGH domain may play a role in binding a moiety in proximity with peptidoglycan, such as a hydrophobic outer membrane lipid or lipopolysaccharide

Amino Acid Patterns around Disulfide Bonds

Disulfide bonds provide an inexhaustible source of information on molecular evolution and biological specificity. In this work, we described the amino acid composition around disulfide bonds in a set of disulfide-rich proteins using appropriate descriptors, based on ANOVA (for all twenty natural amino acids or classes of amino acids clustered according to their chemical similarities) and Scheffé (for the disulfide-rich proteins superfamilies) statistics. We found that weakly hydrophilic and aromatic amino acids are quite abundant in the regions around disulfide bonds, contrary to aliphatic and hydrophobic amino acids. The density distributions (as a function of the distance to the center of the disulfide bonds) for all defined entities presented an overall unimodal behavior: the densities are null at short distances, have maxima at intermediate distances and decrease for long distances. In the end, the amino acid environment around the disulfide bonds was found to be different for different superfamilies, allowing the clustering of proteins in a biologically relevant way, suggesting that this type of chemical information might be used as a tool to assess the relationship between very divergent sets of disulfide-rich proteins

Hyperdimensional Analysis of Amino Acid Pair Distributions in Proteins

Our manuscript presents a novel approach to protein structure analyses. We have organized an 8-dimensional data cube with protein 3D-structural information from 8706 high-resolution non-redundant protein-chains with the aim of identifying packing rules at the amino acid pair level. The cube contains information about amino acid type, solvent accessibility, spatial and sequence distance, secondary structure and sequence length. We are able to pose structural queries to the data cube using program ProPack. The response is a 1, 2 or 3D graph. Whereas the response is of a statistical nature, the user can obtain an instant list of all PDB-structures where such pair is found. The user may select a particular structure, which is displayed highlighting the pair in question. The user may pose millions of different queries and for each one he will receive the answer in a few seconds. In order to demonstrate the capabilities of the data cube as well as the programs, we have selected well known structural features, disulphide bridges and salt bridges, where we illustrate how the queries are posed, and how answers are given. Motifs involving cysteines such as disulphide bridges, zinc-fingers and iron-sulfur clusters are clearly identified and differentiated. ProPack also reveals that whereas pairs of Lys residues virtually never appear in close spatial proximity, pairs of Arg are abundant and appear at close spatial distance, contrasting the belief that electrostatic repulsion would prevent this juxtaposition and that Arg-Lys is perceived as a conservative mutation. The presented programs can find and visualize novel packing preferences in proteins structures allowing the user to unravel correlations between pairs of amino acids. The new tools allow the user to view statistical information and visualize instantly the structures that underpin the statistical information, which is far from trivial with most other SW tools for protein structure analysis

Bioinformatic studies of small disulphide-rich proteins (SDPs)

Ph.DDOCTOR OF PHILOSOPH

LinkProt : a database collecting information about biological links

Protein chains are known to fold into topologically complex shapes, such as knots, slipknots or complex lassos. This complex topology of the chain can be considered as an additional feature of a protein, separate from secondary and tertiary structures. Moreover, the complex topology can be defined also as one additional structural level. The LinkProt database (http://linkprot.cent.uw.edu.pl) collects and displays information about protein links - topologically non-trivial structures made by up to four chains and complexes of chains (e.g. in capsids). The database presents deterministic links (with loops closed, e.g. by two disulfide bonds), links formed probabilistically and macromolecular links. The structures are classified according to their topology and presented using the minimal surface area method. The database is also equipped with basic tools which allow users to analyze the topology of arbitrary (bio)polymers

Oxidative Folding of a Cystine Knot Protein: the Amaranthus α-amylase Inhibitor

The work described in this thesis is concerned with the study of the oxidative folding pathway of a cystine knot protein, Amaranthus a-amylase inhibitor (AAI). The data presented in the thesis are obtained with a combination of different biochemical and biophysical techniques, with an emphasis on NMR spectroscopy. Chapter 1 gives a short introduction to the structural and functional superfamilies of the protein in question. Additionally, it outlines the protein folding problem, with an emphasis on oxidative folding, and stresses the importance of studying such pathways in small proteins. Chapter 2 covers both the basis of experimental techniques and the details of experimental methods used in the study. The remaining four chapters (3, 4, 5 and 6) are results and discussion chapters. Each one deals with a specific aspect of the study of the oxidative folding pathway of Amaranthus a-amylase inhibitor. Chapter 3 presents the results from an improved solid phase peptide synthesis approach, which increased the overall yield of peptide synthesis and recovered a greater amount of AAI after the refolding step. In Chapter 4 the intermediate species that appear on the oxidative folding pathway are identified and characterised in terms of their disulfide content. An illustration of the structural features of the main folding intermediate (MFI) with NMR, photo-CID NP and computer modelling techniques is given in Chapter 5. In Chapter 6 the kinetic process of the oxidative folding of Amaranthus a-amylase inhibitor is studied at the level of a single disulfide species and at the level of a single amino acid residue. The novelty of this thesis is two-fold. First, from the point of view of biological interest, this is the first time that oxidative folding intermediates with non-native disulfide bridges between adjacent cysteine residues are observed. Second, from the point of view of development of biophysical methods, the first application of time-resolved NMR and photo-CIDNP spectroscopy to a complete oxidative folding process is described

Role of accelerated segment switch in exons to alter targeting (ASSET) in the molecular evolution of snake venom proteins

<p>Abstract</p> <p>Background</p> <p>Snake venom toxins evolve more rapidly than other proteins through accelerated changes in the protein coding regions. Previously we have shown that accelerated segment switch in exons to alter targeting (ASSET) might play an important role in its functional evolution of viperid three-finger toxins. In this phenomenon, short sequences in exons are radically changed to unrelated sequences and hence affect the folding and functional properties of the toxins.</p> <p>Results</p> <p>Here we analyzed other snake venom protein families to elucidate the role of ASSET in their functional evolution. ASSET appears to be involved in the functional evolution of three-finger toxins to a greater extent than in several other venom protein families. ASSET leads to replacement of some of the critical amino acid residues that affect the biological function in three-finger toxins as well as change the conformation of the loop that is involved in binding to specific target sites.</p> <p>Conclusion</p> <p>ASSET could lead to novel functions in snake venom proteins. Among snake venom serine proteases, ASSET contributes to changes in three surface segments. One of these segments near the substrate binding region is known to affect substrate specificity, and its exchange may have significant implications for differences in isoform catalytic activity on specific target protein substrates. ASSET therefore plays an important role in functional diversification of snake venom proteins, in addition to accelerated point mutations in the protein coding regions. Accelerated point mutations lead to fine-tuning of target specificity, whereas ASSET leads to large-scale replacement of multiple functionally important residues, resulting in change or gain of functions.</p