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Sequence Database Setup: MSIPI

SwissProt Release 56.0

MSIPI includes a number of SwissProt entries representing common contaminants. As a result, it was affected by the change in the syntax of the Fasta title line in SwissProt release 56.0 (UniProtKB 14.0). This page has been updated to illustrate the new parse rules.

Overview

MSIPI is a database derived from IPI that contains additional information about cSNPs, N-terminus peptides, and known variants in a format suitable for mass spectrometry search engines. MSIPI is produced by the Max-Planck Institute for Biochemistry at Martinsried and the University of Southern Denmark, and distributed by the EBI.

There are currently two MSIPI databases, Human and Mouse. This document uses the Human database as an example. To work with the Mouse database, simply substitute the word "MOUSE" for "HUMAN".

Download

ftp://ftp.ebi.ac.uk/pub/databases/IPI/msipi/current/ for the latest release.
ftp://ftp.ebi.ac.uk/pub/databases/IPI/msipi/old/ for earlier releases.

There are two files: a Fasta database file, (msipi.HUMAN_slim.fasta.gz), and file that also includes the reversed sequences, (msipi.HUMAN_decoy_slim.fasta.gz), providing a ready-made, concatenated decoy database. The inclusion of the word slim in the file name indicates that the title has been abbreviated.

There isn't an easy way to configure a local full text reference file because MSIPI has a number of Swiss-Prot sequences appended to the end, representing common contaminants. Mascot requires a 1:1 correspondence between the Fasta file and the full text reference file, so the standard IPI *.dat file cannot be used. Fortunately, you can query an SRS server to get full text reports, as shown in the configuration example below.

Use of code letter J

As described in the Nature Methods paper, the additional peptide sequences in MSIPI, representing cSNPs, N-terminus peptides, and known variants, are delimited from the original protein sequences using the letter J. This code letter is not used in a default Mascot configuration, and has a mass of zero. You need to add this letter to any enzyme definitions that you plan to use with MSIPI as an additional cleavage site. To avoid confusion, create new definitions with self-explanatory names, such as TrypsinMSIPI.

Mascot Configuration Editor

For earlier versions of Mascot, use a text editor to modify the enzymes configuration file:

Title:TrypsinMSIPI Cleavage[0]:KR Restrict[0]:P Cterm[0] Cleavage[1]:J Cterm[1] Cleavage[2]:J Nterm[2] * Title:TrypsinMSIPI/P Cleavage[0]:KRJ Cterm[0] Cleavage[1]:J Nterm[1] *

The supplementary material to the Nature Methods paper suggests setting the mass of J to that of Asparagine, (114). However, the letter J is sometimes used as the ambiguity code for I or L. If you search databases in which J is used in this way, you will need to set the mass of J to 113. This will mean that, when searching MSIPI with missed cleavages set greater than zero, you will sometimes see spurious peptide sequences containing J among the low scoring, random matches.

Otherwise, best to leave the mass of J at zero. You will still see peptide sequences containing J in the result reports, but this will not change any of the mass values or the matches.

Taxonomy

Taxonomy is not required because all entries are from the same species

Parse Rules

Typical Fasta title lines from the non-decoy Fasta files are:

>MSIPI:IPI00000001.2| Gene_Symbol=STAU1 Isoform Long of Double-stranded RNA-binding protein Staufen homolog 1 lng=577 # CON[595,R,359,A] #

>MSIPI:sp|P07758|A1AT1_MOUSE Alpha-1-antitrypsin 1-1 OS=Mus musculus GN=Serpina1a PE=1 SV=4

The parse rule must be general enough to match both the IPI entries and the Swiss-Prot contaminant entries.

Accession from Fasta title: ">MSIPI:s*p*|*\([^| .]*\)"
Description from Fasta title: ">[^ ]* \(.*\)"

These rules will not work for the Fasta file that includes concatenated decoy entries. It doesn't appear possible for a single parse rule to extract accession strings for the target entries that will be recognised by an SRS server and still extract unique accessions for the reversed SwissProt sequences.

The following rule will extract a unique accession string, but the link for Protein View will fail:

Accession from Fasta title: ">MSIPI:\([^ .]*\)"

Configuration

For this example, the Fasta file was downloaded to C:\sequence\MSIPI_human\current, decompressed using gzip, and renamed to MSIPI_human_3.48.fasta.

Mascot database maintenance utility

As mentioned earlier, there isn't a local reference file corresponding to the Fasta. Full text for individual entries can be retrieved from an SRS server, such as the public server at EBI. The following syntax for the Path field retrieves entries from both IPI and Swiss-Prot:

/srsbin/cgi-bin/wgetz?-e+[{IPI%20SWISSPROT}-acc:#ACCESSION#]+-vn+2

Mascot database maintenance utility

If you get the message Error: Nothing returned by call to ... instead of the sample full text report, this usually indicates that the Mascot Server cannot access the Internet because of a firewall or proxy server problem. In the case of a proxy server, the host, port and, if necessary, a user name and password are defined in mascot.dat using the labels proxy_server, proxy_username, and proxy_password. Choose Edit Options in Database Maintenance. For example:

proxy_server http://my_proxy:3128

If you don't require full text in a Mascot Protein View report, simply leave the Host, Port, and Path fields blank and choose
--- no full text report ---
in the drop down list.

Always test a new definition before applying the changes to mascot.dat.