Sequence search algorithm assessment and testing toolkit (SAT)

MOTIVATION: The Sequence Search Algorithm Assessment and Testing Toolkit (SAT) aims to be a complete package for the comparison of different protein homology search algorithms. The structural classification of proteins can provide us with a clear criterion for judgment in homology detection. There have been several assessments based on structural sequences with classifications but a good deal of similar work is now being repeated with locally developed procedures and programs. The SAT will provide developers with a complete package which will save time and produce more comparable performance assessments for search algorithms. The package is complete in the sense that it provides a non-redundant large sequence resource database, a well-characterized query database of proteins domains, all the parsers and some previous results from PSI-BLAST and a hidden markov model algorithm. RESULTS: An analysis on two different data sets was carried out using the SAT package. It compared the performance of a full protein sequence database (RSDB100) with a non-redundant representative sequence database derived from it (RSDB50). The performance measurement indicated that the full database is sub-optimal for a homology search. This result justifies the use of much smaller and faster RSDB50 than RSDB100 for the SAT. AVAILABILITY: A web site is up. The whole packa ge is accessible via www and ftp. ftp://ftp.ebi.ac.uk/pub/contrib/jong/SAT http://cyrah.ebi.ac.uk:1111/Proj/Bio/SAT http://www.mrc-lmb.cam.ac.uk/genomes/SAT In the package, some previous assessment results produced by the package can also be found for reference. CONTACT: jong@ebi.ac.uk     

EMBL-Align: a new public nucleotide and amino acid multiple sequence alignment database

The submission of multiple sequence alignment data to EMBL has grown 30-fold in the past 10 years, creating a problem of archiving them. The EBI has developed a new public database of multiple sequence alignments called EMBL-Align. It has a dedicated web-based submission tool, Webin-Align. Together they represent a comprehensive data management solution for alignment data. Webin-Align accepts all the common alignment formats and can display data in CLUSTALW format as well as a new standard EMBL-Align flat file format. The alignments are stored in the EMBL-Align database and can be queried from the EBI SRS (Sequence Retrieval System) server. AVAILABILITY: Webin-Align: http://www.ebi.ac.uk/embl/Submission/align_top.html, EMBL-Align: ftp://ftp.ebi.ac.uk/pub/databases/embl/align, http://srs.ebi.ac.uk/     

The EMBL nucleotide sequence database

The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl/) is maintained at the European Bioinformatics Institute (EBI) in an international collaboration with the DNA Data Bank of Japan (DDBJ) and GenBank at the NCBI (USA). Data is exchanged amongst the collaborating databases on a daily basis. The major contributors to the EMBL database are individual authors and genome project groups. Webin is the preferred web-based submission system for individual submitters, whilst automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO). Database releases are produced quarterly. Network services allow free access to the most up-to-date data collection via ftp, email and World Wide Web interfaces. EBI's Sequence Retrieval System (SRS), a network browser for databanks in molecular biology, integrates and links the main nucleotide and protein databases plus many specialized databases. For sequence similarity searching a variety of tools (e.g. Blitz, Fasta, BLAST) are available which allow external users to compare their own sequences against the latest data in the EMBL Nucleotide Sequence Database and SWISS-PROT.     

UniRef: comprehensive and non-redundant UniProt reference clusters

MOTIVATION: Redundant protein sequences in biological databases hinder sequence similarity searches and make interpretation of search results difficult. Clustering of protein sequence space based on sequence similarity helps organize all sequences into manageable datasets and reduces sampling bias and overrepresentation of sequences. RESULTS: The UniRef (UniProt Reference Clusters) provide clustered sets of sequences from the UniProt Knowledgebase (UniProtKB) and selected UniProt Archive records to obtain complete coverage of sequence space at several resolutions while hiding redundant sequences. Currently covering >4 million source sequences, the UniRef100 database combines identical sequences and subfragments from any source organism into a single UniRef entry. UniRef90 and UniRef50 are built by clustering UniRef100 sequences at the 90 or 50% sequence identity levels. UniRef100, UniRef90 and UniRef50 yield a database size reduction of approximately 10, 40 and 70%, respectively, from the source sequence set. The reduced redundancy increases the speed of similarity searches and improves detection of distant relationships. UniRef entries contain summary cluster and membership information, including the sequence of a representative protein, member count and common taxonomy of the cluster, the accession numbers of all the merged entries and links to rich functional annotation in UniProtKB to facilitate biological discovery. UniRef has already been applied to broad research areas ranging from genome annotation to proteomics data analysis. AVAILABILITY: UniRef is updated biweekly and is available for online search and retrieval at http://www.uniprot.org, as well as for download at ftp://ftp.uniprot.org/pub/databases/uniprot/uniref. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.     

The InterPro Database, 2003 brings increased coverage and new features

InterPro, an integrated documentation resource of protein families, domains and functional sites, was created in 1999 as a means of amalgamating the major protein signature databases into one comprehensive resource. PROSITE, Pfam, PRINTS, ProDom, SMART and TIGRFAMs have been manually integrated and curated and are available in InterPro for text- and sequence-based searching. The results are provided in a single format that rationalises the results that would be obtained by searching the member databases individually. The latest release of InterPro contains 5629 entries describing 4280 families, 1239 domains, 95 repeats and 15 post-translational modifications. Currently, the combined signatures in InterPro cover more than 74% of all proteins in SWISS-PROT and TrEMBL, an increase of nearly 15% since the inception of InterPro. New features of the database include improved searching capabilities and enhanced graphical user interfaces for visualisation of the data. The database is available via a webserver (http://www.ebi.ac.uk/interpro) and anonymous FTP (ftp://ftp.ebi.ac.uk/pub/databases/interpro).     

InterProScan--an integration platform for the signature-recognition methods in InterPro

InterProScan is a tool that scans given protein sequences against the protein signatures of the InterPro member databases, currently--PROSITE, PRINTS, Pfam, ProDom and SMART. The number of signature databases and their associated scanning tools as well as the further refinement procedures make the problem complex. InterProScan is designed to be a scalable and extensible system with a robust internal architecture. AVAILABILITY: The Perl-based InterProScan implementation is available from the EBI ftp server (ftp://ftp.ebi.ac.uk/pub/software/unix/iprscan/) and the SRS-basedInterProScan is available upon request. We provide the public web interface (http://www.ebi.ac.uk/interpro/scan.html) as well as email submission server (interproscan@ebi.ac.uk).     

Fast assignment of protein structures to sequences using the intermediate sequence library PDB-ISL

MOTIVATION: For large-scale structural assignment to sequences, as in computational structural genomics, a fast yet sensitive sequence search procedure is essential. A new approach using intermediate sequences was tested as a shortcut to iterative multiple sequence search methods such as PSI-BLAST. RESULTS: A library containing potential intermediate sequences for proteins of known structure (PDB-ISL) was constructed. The sequences in the library were collected from a large sequence database using the sequences of the domains of proteins of known structure as the query sequences and the program PSI-BLAST. Sequences of proteins of unknown structure can be matched to distantly related proteins of known structure by using pairwise sequence comparison methods to find homologues in PDB-ISL. Searches of PDB-ISL were calibrated, and the number of correct matches found at a given error rate was the same as that found by PSI-BLAST. The advantage of this library is that it uses pairwise sequence comparison methods, such as FASTA or BLAST2, and can, therefore, be searched easily and, in many cases, much more quickly than an iterative multiple sequence comparison method. The procedure is roughly 20 times faster than PSI-BLAST for small genomes and several hundred times for large genomes. AVAILABILITY: Sequences can be submitted to the PDB-ISL servers at http://stash.mrc-lmb.cam.ac.uk/PDB_ISL/ or http://cyrah.ebi.ac.uk:1111/Serv/PDB_ISL/ and can be downloaded from ftp://ftp.ebi.ac.uk/pub/contrib/jong/PDB_+ ++ISL/ CONTACT: sat@mrc-lmb.cam.ac.uk and jong@ebi.ac.uk     

KIND-a non-redundant protein database

SUMMARY: KIND (Karolinska Institutet Nonredundant Database) is a protein database where identical sequences, both full length and partial, have been removed. The database contains nearly 274 900 sequences, half of which originate from the protein sequence databases Swissprot and PIR, while the other half come from translated open reading frames in GenPept and TrEMBL. AVAILABILITY: KIND is downloadable from ftp://ftp.mbb.ki.se/pub/KIND.     

VARSPLIC: alternatively-spliced protein sequences derived from SWISS-PROT and TrEMBL

SUMMARY: The program varsplic.pl uses information present in the SWISS-PROT and TrEMBL databases to create new records for alternatively spliced isoforms. These new records can be used in similarity searches. AVAILABILITY: The program is available at ftp://ftp.ebi.ac.uk/pub/software/swissprot/, together with regularly updated output files. CONTACT: pkersey@ebi.ac.uk     

The EMBL Nucleotide Sequence Database

The EMBL Nucleotide Sequence Database (aka EMBL-Bank; http://www.ebi.ac.uk/embl/) incorporates, organises and distributes nucleotide sequences from all available public sources. EMBL-Bank is located and maintained at the European Bioinformatics Institute (EBI) near Cambridge, UK. In an international collaboration with DDBJ (Japan) and GenBank (USA), data are exchanged amongst the collaborating databases on a daily basis. Major contributors to the EMBL database are individual scientists and genome project groups. Webin is the preferred web-based submission system for individual submitters, whilst automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO). Database releases are produced quarterly. Network services allow free access to the most up-to-date data collection via FTP, email and World Wide Web interfaces. EBI’s Sequence Retrieval System (SRS), a network browser for databanks in molecular biology, integrates and links the main nucleotide and protein databases plus many other specialized databases. For sequence similarity searching, a variety of tools (e.g. Blitz, Fasta, BLAST) are available which allow external users to compare their own sequences against the latest data in the EMBL Nucleotide Sequence Database and SWISS-PROT. All resources can be accessed via the EBI home page at http://www.ebi.ac.uk.     

Exhaustive enumeration of protein domain families

Domains are considered as the basic units of protein folding, evolution, and function. Decomposing each protein into modular domains is thus a basic prerequisite for accurate functional classification of biological molecules. Here, we present ADDA, an automatic algorithm for domain decomposition and clustering of all protein domain families. We use alignments derived from an all-on-all sequence comparison to define domains within protein sequences based on a global maximum likelihood model. In all, 90% of domain boundaries are predicted within 10% of domain size when compared with the manual domain definitions given in the SCOP database. A representative database of 249,264 protein sequences were decomposed into 450,462 domains. These domains were clustered on the basis of sequence similarities into 33,879 domain families containing at least two members with less than 40% sequence identity. Validation against family definitions in the manually curated databases SCOP and PFAM indicates almost perfect unification of various large domain families while contamination by unrelated sequences remains at a low level. The global survey of protein-domain space by ADDA confirms that most large and universal domain families are already described in PFAM and/or SMART. However, a survey of the complete set of mobile modules leads to the identification of 1479 new interesting domain families which shuffle around in multi-domain proteins. The data are publicly available at ftp://ftp.ebi.ac.uk/pub/contrib/heger/adda.     

PROF_ PAT 1.3: updated database of patterns used to detect local similarities

MOTIVATION: When analysing novel protein sequences, it is now essential to extend search strategies to include a range of 'secondary' databases. Pattern databases have become vital tools for identifying distant relationships in sequences, and hence for predicting protein function and structure. The main drawback of such methods is the relatively small representation of proteins in trial samples at the time of their construction. Therefore, a negative result of an amino acid sequence comparison with such a databank forces a researcher to search for similarities in the original protein banks. We developed a database of patterns constructed for groups of related proteins with maximum representation of amino acid sequences of SWISS-PROT in the groups. RESULTS: Software tools and a new method have been designed to construct patterns of protein families. By using such method, a new version of databank of protein family patterns, PROF_ PAT 1.3, is produced. This bank is based on SWISS-PROT (r1.38) and TrEMBL (r1.11), and contains patterns of more than 13 000 groups of related proteins in a format similar to that of the PROSITE. Motifs of patterns, which had the minimum level of probability to be found in random sequences, were selected. Flexible fast search program accompanies the bank. The researcher can specify a similarity matrix (the type PAM, BLOSUM and other). Variable levels of similarity can be set (permitting search strategies ranging from exact matches to increasing levels of 'fuzziness'). AVAILABILITY: The Internet address for comparing sequences with the bank is: http://wwwmgs.bionet.nsc.ru/mgs/programs/prof_pat/. The local version of the bank and search programs (approximately 50 Mb) is available via ftp: ftp://ftp.bionet.nsc. ru/pub/biology/vector/prof_pat/, and ftp://ftp.ebi.ac. uk/pub/databases/prof_pat/. Another appropriate way for its external use is to mail amino acid sequences to bachin@vector.nsc.ru for comparison with PROF_ PAT 1.3.     

Post-processing of BLAST results using databases of clustered sequences

Motivation: When evaluating the results of a sequence similaritysearch, there are many situations where it can be useful todetermine whether sequences appearing in the results share somedistinguishing characteristic. Such dependencies between databaseentries are often not readily identifiable, but can yield importantnew insights into the biological function of a gene or protein. Results: We have developed a program called CBLAST that sortsthe results of a BLAST sequence similarity search accordingto sequence membership in user-defined ‘clusters’of sequences. To demonstrate the utility of this application,we have constructed two cluster databases. The first describesclusters of nucleotide sequences representing the same gene,as documented in the UNIGENE database, and the second describesclusters of protein sequences which are members of the proteinfamilies documented in the PROSITE database. Cluster databasesand the CBLAST post-processor provide an efficient mechanismfor identifying and exploring relationships and dependenciesbetween new sequences and database entries. Availability: The software described in this article is availablefree of charge from the EBI software archive at < ftp: //ftp.ebi. ac. uk/pub/software/unix >. Contact: E-mail: rainer _fuchs@glaxowellcome.com     

The peptaibol database: a sequence and structure resource.

The peptaibols are a large family of membrane-active peptides with considerable sequence homology, but with different biological properties and three-dimensional structures. They constitute a rich resource of naturally occurring 'mutants' which are potentially valuable for structure/function studies of ion channels. A searchable on-line database of sequences and structures of the peptaibols has been created at http://www.cryst.bbk.ac.uk/peptaibol, as a resource for the biological and structural community. In this paper, the contents and organization of the website are discussed as well as procedures for submission of new entries to the database. At present, more than 300 peptaibol sequences are stored in the database. Each sequence entry contains its full literature reference and information about its biological source. Tools are provided for searching for specific peptaibol sequences or groupings of sequences, and for locating peptaibols containing specified sequence motifs. In addition the website acts as a database for structural information. The coordinates of all currently available peptaibol x-ray and NMR structures are included and complemented, where appropriate. with molecular graphics illustrations. These include figures of model channel structures and comparisons between different peptaibol structures. The peptaibol database thus provides a tool for ready access to information and a means of investigating the sequences and structures of this class of polypeptides.     

A RAPID algorithm for sequence database comparisons: application to the identification of vector contamination in the EMBL databases

MOTIVATION: Word-matching algorithms such as BLAST are routinely used for sequence comparison. These algorithms typically use areas of matching words to seed alignments which are then used to assess the degree of sequence similarity. In this paper, we show that by formally separating the word-matching and sequence-alignment process, and using information about word frequencies to generate alignments and similarity scores, we can create a new sequence-comparison algorithm which is both fast and sensitive. The formal split between word searching and alignment allows users to select an appropriate alignment method without affecting the underlying similarity search. The algorithm has been used to develop software for identifying entries in DNA sequence databases which are contaminated with vector sequence. RESULTS: We present three algorithms, RAPID, PHAT and SPLAT, which together allow vector contaminations to be found and assessed extremely rapidly. RAPID is a word search algorithm which uses probabilities to modify the significance attached to different words; PHAT and SPLAT are alignment algorithms. An initial implementation has been shown to be approximately an order of magnitude faster than BLAST. The formal split between word searching and alignment not only offers considerable gains in performance, but also allows alignment generation to be viewed as a user interface problem, allowing the most useful output method to be selected without affecting the underlying similarity search. Receiver Operator Characteristic (ROC) analysis of an artificial test set allows the optimal score threshold for identifying vector contamination to be determined. ROC curves were also used to determine the optimum word size (nine) for finding vector contamination. An analysis of the entire expressed sequence tag (EST) subset of EMBL found a contamination rate of 0.27%. A more detailed analysis of the 50 000 ESTs in est10.dat (an EST subset of EMBL) finds an error rate of 0.86%, principally due to two large-scale projects. AVAILABILITY: A Web page for the software exists at http://bioinf.man.ac.uk/rapid, or it can be downloaded from ftp://ftp.bioinf.man.ac.uk/RAPID CONTACT: crispin@cs.man.ac.uk     

HOMSTRAD: adding sequence information to structure-based alignments of homologous protein families

summary: We describe an extension to the Homologous Structure Alignment Database (HOMSTRAD; Mizuguchi et al., Protein Sci., 7, 2469-2471, 1998a) to include homologous sequences derived from the protein families database Pfam (Bateman et al., Nucleic Acids Res., 28, 263-266, 2000). HOMSTRAD is integrated with the server FUGUE (Shi et al., submitted, 2001) for recognition and alignment of homologues, benefitting from the combination of abundant sequence information and accurate structure-based alignments. AVAILABILITY The HOMSTRAD database is available at: http://www-cryst.bioc.cam.ac.uk/homstrad/. Query sequences can be submitted to the homology recognition/alignment server FUGUE at: http://www-cryst.bioc.cam.ac.uk/fugue/.     

iPfam: visualization of protein-protein interactions in PDB at domain and amino acid resolutions

SUMMARY: There are many resources that contain information about binary interactions between proteins. However, protein interactions are defined by only a subset of residues in any protein. We have implemented a web resource that allows the investigation of protein interactions in the Protein Data Bank structures at the level of Pfam domains and amino acid residues. This detailed knowledge relies on the fact that there are a large number of multidomain proteins and protein complexes being deposited in the structure databases. The resource called iPfam is hosted within the Pfam UK website. Most resources focus on the interactions between proteins; iPfam includes these as well as interactions between domains in a single protein. AVAILABILITY: iPfam is available on the Web for browsing at http://www.sanger.ac.uk/Software/Pfam/iPfam/; the source-data for iPfam is freely available in relational tables via the ftp site ftp://ftp.sanger.ac.uk/pub/databases/Pfam/database_files/.     

The EMBL nucleotide sequence database

The European Molecular Biology Laboratory (EMBL) Nucleotide Sequence Database (http://www.ebi.ac. uk/embl/index.html ) is maintained at the European Bioinformatics Institute (EBI) in an international collaboration with the DNA Data Bank of Japan (DDBJ) and GenBank (USA). Data is exchanged amongst the collaborative databases on a daily basis. The major contributors to the EMBL database are individual authors and genome project groups. WEBIN is the preferred web-based submission system for individual submitters, whilst automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO). Database releases are produced quarterly. Network services allow free access to the most up-to-date data collection via Internet and WWW interfaces. EBI's Sequence Retrieval System (SRS) is a network browser for databanks in molecular biology, integrating and linking the main nucleotide and protein databases plus many specialised databases. For sequence similarity searching a variety of tools (e.g., BLITZ, FASTA, BLAST) are available which allow external users to compare their own sequences against the most currently available data in the EMBL Nucleotide Sequence Database and SWISS-PROT.     

Vertebrate MitBASE: a specialised database on vertebrate mitochondrial DNA sequences.

Vertebrate MitBASE is a specialized database where all the vertebrate mitochondrial DNA entries from primary databases are collected, revised and integrated with new information emerging from the literature. Variant sequences are also analyzed, aligned and linked to reference sequences. Data related to the same species and fragment can be viewed over the WWW. The database has a flexible interface and a retrieval system to help non-expert users and contains information not currently available in the primary databases. Vertebrate MitBASE is now available through the MitBASE home page at URL: http://www.ebi.ac.uk/htbin/Mitbase/mitb ase.pl. This work is part of a larger project, MitBASE which is a network of databases covering the full panorama of knowledge on mitochondrial DNA from protists to human sequences.     

Clustal W and Clustal X version 2.0

SUMMARY: The Clustal W and Clustal X multiple sequence alignment programs have been completely rewritten in C++. This will facilitate the further development of the alignment algorithms in the future and has allowed proper porting of the programs to the latest versions of Linux, Macintosh and Windows operating systems. AVAILABILITY: The programs can be run on-line from the EBI web server: http://www.ebi.ac.uk/tools/clustalw2. The source code and executables for Windows, Linux and Macintosh computers are available from the EBI ftp site ftp://ftp.ebi.ac.uk/pub/software/clustalw2/