PSIPRED RELEASE NOTES=====================PSIPRED Version 2.45By David Jones, November 2003Here are some very brief notes on using the PSIPRED V2 software.PSIPRED is supplied in source code form - it must be compiled beforeit can be used. The code should compile on any ANSI C compiler e.g.the GNU C compiler.Please see the LICENSE file for the license terms for the software.Basically it's free to anyone (including commercial users) as long asyou don't want to sell the software or, for example, store the resultsobtained with it in a database and then try to sell the database.If you do wish to sell the software or use it in a commercial product,then please contact Inpharmatica Ltd. (http://www.inpharmatica.co.uk).PSIPRED is run via a tcsh shell script called "runpsipred" - this is avery simple script which you should be able to convert to Perl or whateverscripting language you like.If your sequence does not have any homologues in the current data banks,then it is possible to run PSIPRED on a single sequence. In this case,PSIPRED is run via a tcsh shell script called "runpsi_single". Unfortunately,like every other secondary structure prediction method, PSIPRED does notperform well on single sequences. Any secondary structure prediction based ona single sequence should be considered as unreliable.Before running PSIPRED, please check the runpsipred and runpsi_single scriptsto see if the path variables are set to wherever you have installed theprogram and data files. The default is to assume that the program isinstalled in the current directory - this is probably NOT what you want!INSTALLATION============Firstly compile the software:tcsh% cd to-wherever-you-untarred-PSIPREDtcsh% cd srctcsh% maketcsh% make installThe executables will be placed in the PSIPRED bin directory.You must also install the PSI-BLAST and Impala software from theNCBI toolkit, and also install appropriate sequence data banks.The NCBI toolkit can be obtained from URL ftp://ftp.ncbi.nih.govPSI-BLAST executables can be obtained from ftp://ftp.ncbi.nih.gov/blastEXAMPLE USAGE=============In this example the target sequence is called "example.fasta":tcsh% runpsipred example.fastaRunning PSI-BLAST with sequence example.fasta ...Predicting secondary structure...Pass1 ...Pass2 ...Cleaning up ...Final output file: example.horizFinished.That's it - you can then look at the output:tcsh% more example.horizSPECIAL OPTIONS===============The psipass2 program has several special options which you can use if you wish.For example, the default command is as follows:psipass2 weights_p2.dat 1 1.0 1.0 output.ss2 input.ss > output.horizArguments 2,3 & 4 are as follows:Argument 2: No of filter iterationsThis controls the amount of "smoothing" that is carried out on the finalprediction. The recommended setting is 1, but it may be worth tryinghigher values to increase the level of smoothing.Argument 3&4: Helix/Strand Decision constantsThese options control the bias for helix (Arg3) and strand (Arg4) predictions.The default values are equal to 1.0, but if you know your protein is, forexample, mostly comprised of beta strands then you can increase the biastowards beta strand prediction. For example:psipass2 weights_p2.dat 1 1.0 1.3 output.ss2 input.ss > output.horizincreases the bias towards beta strand prediction by approximately 30%. SEQUENCE DATA BANK==================It is important to ensure than the sequence data bank used with PSI-BLASThas been filtered to remove low-complexity regions, transmembrane regions,and coiled-coil segments. If this is not done, then it is essential thatthe PSI-BLAST output for the target sequence is checked by-eye to ensurethat no spurious sequences have been included in the PSI-BLAST alignment.A program called "pfilt" is included which will filter FASTA files beforeusing the formatdb command to generate the encoded BLAST data bank files.For example:tcsh% pfilt nr.fasta > filtnrtcsh% formatdb -t filtnr -i filtnrtcsh% cp filtnr.p?? $BLASTDB(note that the above command assumes you have already set the BLASTDBenvironment variable to the directory where you usually keep yourBLAST data banks)CHANGES FROM THE ORIGINAL PSIPRED=================================The following is a quick summary of the main changes since the originalPSIPRED.1. The program now makes use of PSI-BLAST binary checkpoint files (using theImpala program makemat) to reduce loss of precision when parsing the originalASCII position specific matrices.2. By default the 1st pass uses an average of 4 different neural networkweight sets - this improves prediction accuracy slightly.3. In addition to the normal horizontal summary output format, the programnow also produces a full table of results which shows the individualcoil, helix, strand network outputs.4. A one-line header is output at the start of the output files to allowTHREADER 3 (and other programs) to automatically recognise a PSIPREDprediction.