This section explains how to open sequences and alignments in CINEMA, both from your own computer and from remote databases.
Opening locally-stored sequences
To open a sequence or alignment file stored locally on your computer, choose Open from the File menu, or click on the open icon on the toolbar. The Open file dialog will appear. If necessary, use the dialog to locate the folder containing the desired sequence file. At present, CINEMA can read alignments and sequences in UTOPIA, c5a (CINEMA 5), FastA, NBRF-PIR or PDB formats.
Sequence files containing more than one chain
Some sequence formats (e.g. PDB) can contain sequence data representing more than one chain. When a sequence with multiple chains is opened in CINEMA, you will be shown a list of the available chain IDs and asked to select one; this is the chain that will be shown for the sequence within the alignment. The choice is not permanent though - the chain can be switched later using the sequence context menu (unless a motif has been defined that covers the sequence.) When an alignment is saved, the chain preferences for any multi-chain sequences are stored, and will be automatically recovered next time the alignment is opened.
Adding new sequences to an alignment
Once you have opened a new alignment, you can add further sequences to it at any time. To add a local sequence file to the current alignment, choose Add local sequence to alignment... from the File menu. An open file dialog will appear; choose an appropriate sequence file and click on the Open button. The chosen sequence will be appended to the end of the alignment.
Saving CINEMA alignments
To save an alignment, choose Save or Save as... from the File menu, or click on the appropriate icon on the toolbar. If you choose Save as... (or if this is the first time a new alignment has been saved) the Save As dialog will appear and you will be prompted to supply a name for the alignment file. If necessary, use the dialog to locate the folder where the alignment file is to be saved, enter a name and click Save.
Sequences can be opened directly in CINEMA from a number of remote databases; currently CINEMA provides direct access to the UniProt (Swiss-Prot & TrEMBL), PDB and UTOPIA-PFF databases.
A search tool, currently known as "Find-O-Matic", allows sequences to be found using their database identifier (e.g. PDB code or UniProt accession), or via a simple free-text search (e.g. by entering "human".) A simple drag-and-drop mechanism is then used to open sequences in CINEMA. Find-O-Matic also allows basic similarity searches, using BLAST. The Find-O-Matic interface is shown below:
Searching for sequences
To search for, and open sequences from a remote database, first ensure you have a working internet connection, and then choose Open From Remote Database... from the File menu, or click on the Open Remote icon on the toolbar. The Find-O-Matic dialog will appear. Enter a search term (e.g. "3EBX", "OPSD_BOVIN" or "mouse") in the box in the top-right corner of the dialog and press the enter key. There will be a slight delay whilst the search progresses and, if successful, the results will then be displayed in the list below. Each result will give the the database ID of the sequence, along with any descriptive data found.
Valid search terms are PDB codes and UniProt accessions/identifiers, all of which will return results directly from the respective databases. Find-O-Matic will also send every query to the UniProt free-text search service so, for example, a search with PDB code "3EBX" will also return all UniProt entries that are, for whatever reason, connected to that PDB entry. Using more general terms, e.g. "sheep" or "human", will also return results from UniProt, but bear in mind that such searches could take a significant amount of time, and potentially return an enormous number of results! Pressing the "Cancel" button (or the Esc key on your keyboard) at any time will halt the process. The UniProt free text search is presently limited (on the server side) to returning at most 20,000 results. We hope to soon provide our own version of this service with no such limitations.
Note Find-O-Matic relies on a number of remote database services, some hosted on the UTOPIA server and others hosted by the RCSB (http://www.rcsb.org/pdb/home/home.do) and PIR (http://www.pir.uniprot.org). We have no control over the availability of the non-UTOPIA services and this means that, from time-to-time, access to these services may "disappear". In such cases it is likely that these services are only temporarily down; the will usually be accessible again soon, when brought back online by their hosts.
We aim to provide continuous and reliable access to the services hosted on the UTOPIA server. However, our services have only recently become "live", and until we have a better idea of the potential demand for them we cannot guarantee that our server will always be accessible.
We maintain a page (http://utopia.cs.manchester.ac.uk/serviceinfo/) listing the services accessible via UTOPIA tools. This page shows which services are currently available and which (if any) are unavailable. If a service is unavailable, we will attempt to provide details of why, and an estimate of when the service will come back online. Please contact us if you are having difficulties accessing services.
To open one of the sequences found in CINEMA, simply click on and drag the result from the Find-O-Matic window and then drop it on the CINEMA control window. A new alignment will be created and the selected sequence will be added to it. If necessary, the sequence will first be fetched from its database, which may result in a short delay before the sequence is opened in CINEMA. Multiple sequences can be opened at the same time by using the control and/or shift keys to selected the required sequences and then dragging and dropping this selection over to the CINEMA control window. Again, there may be a delay whilst the sequences are fetched from their database(s).
The image below illustrates this behaviour:
Adding sequences to an existing alignment
Sequences found using Find-O-Matic can be added to an existing alignment by dragging them over to and dropping them on the Alignment window. The new sequence(s) will either be inserted into, or appended to the end of the alignment, depending on where it is dropped, as illustrated by a dashed blue line. The image below illustrates this behaviour:
After searching it is then possible to perform a similarity search on one of the results, using BLAST (specifically the blastp algorithm, searching against the Swiss-Prot database.) To perform a BLAST search, right-click on the result you wish to BLAST and choose BLAST xxx for homologues... from the context menu:
After a short delay, Find-O-Matic will list the results of the BLAST search, which can be opened by dragging the required sequence(s) into CINEMA in the manner described above.
Presently, we use the BLAST service provided by DDBJ (http://xml.nig.ac.jp/wsdl/Blast.wsdl), calling the "searchSimple" operation, which performs a protein-protein BLAST against Swiss-Prot and leaves all other parameters at their default values. In future releases, we will provide access to a much greater range of search services.
CINEMA uses the UTOPIA bioinformatics data model and associated file format. In a UTOPIA alignment file, sequences are stored separately from gap insertions, motifs and other annotations. In fact, the sequences themselves may not necessarily be held directly within the alignment file - a CINEMA alignment may hold just references to a set of sequences, e.g. as UniProt database IDs. When alignment file is opened in CINEMA, any external sequences it references are automatically obtained from the appropriate locations (e.g. from a remote database), the gaps are re-inserted and any other annotations defined (e.g. motifs) are reconstructed.
The main advantage of this approach is that it separates axiomatic data (i.e. the sequences) from annotations (e.g. gaps). This means that the sequences are left unadulterated and consistent with the archived versions in the databases (they may actually be the archived versions.)
Alignments created outside of CINEMA (e.g. with other alignment tools or as a hand-compiled text file of sequences) and alignments created with older versions of CINEMA (e.g. .c5a alignments, hereafter referred to as "legacy" alignments) are, unfortunately, not guaranteed to be 100% compatible with the latest release.
CINEMA will happily open non-UTOPIA and legacy alignment files, and will automatically convert them into UTOPIA format, restoring any gaps and motifs that may be present. All standard CINEMA alignment tasks can be performed with such an alignment - visualisation, inserting/removing gaps, defining motifs, etc. However, some more advanced (and future) aspects of CINEMA functionality may not work correctly (e.g collaborative work with the Ambrosia protein structure viewer.) As such, you will be warned whenever you try to open a non-UTOPIA or legacy alignment file. This warning may be turned off by checking the "Don't warn me again" box before selecting Open.
Converting a non-UTOPIA or legacy CINEMA alignment
CINEMA provides an interactive tool, the Legacy Alignment Converter, for converting non-UTOPIA or legacy CINEMA alignments into the new UTOPIA format. Converting an alignment is a user-driven process that involves providing alternative sources for each of the sequences contained in the alignment. For example, if you wish to convert an alignment containing 4 sequences, each originally obtained from Swiss-Prot, you will need to show CINEMA where individual copies of each of these sequences can be found, either as files stored on your computer or by providing the appropriate ID/Accession codes so they may be obtained directly from the UniProt/Swiss-Prot database. If you use sequences stored on you computer then they should be unadulterated, i.e. they should have no gap insertions or other annotations. Once all the sequences have been accounted for, and replacements found, then any annotations attached to the legacy alignment, e.g. gaps, motifs/fingerprints or breakpoints, can be automatically added to the new alignment.
To convert a legacy alignment, choose Convert a legacy alignment... from the File menu. The Legacy Alignment Converter dialog box will appear; click on Open to load the alignment you wish to convert. Each of the sequences found in the alignment will be listed in the centre of the dialog box; click the Start conversion button to begin. You will be asked to provide an alternative for the first sequence found in the alignment - you can choose a locally-stored sequence file, or ask CINEMA to obtain the sequence directly from either the UniProt or PDB databases. This process is then repeated until you have provided alternatives for each of the sequences. If there are any gaps in the legacy alignment, or other annotations CINEMA recognises (such as motifs or breakpoints) then you will be asked if you wish to convert them as well. Finally, you will be asked to save the newly created alignment file. Once the conversion process is complete, you are given the option of opening the newly converted alignment up in CINEMA or of converting another alignment.