MassSorter

a tool for administrating and analyzing data from
mass spectrometry experiments on proteins
with known amino acid sequences

Contents:

• What is MassSorter?
• What does MassSorter do?
• What MassSorter is not
• What kind of data can MassSorter use?
• What kind of data does MassSorter generate?
• Color Coding

What is MassSorter?

MassSorter is a program for administrating and analyzing data from MALDI-TOF mass spectrometry (MS) experiments on proteins with known amino acid sequences.

MassSorter has been developed by Harald Barsnes from the University of Bergen during 2004/2007, after an initial idea by Dr. Svein-Ole Mikalsen from the Norwegian Radium Hospital in Oslo.

Any questions or requests regarding MassSorter can be forwarded to MassSorter@ii.uib.no.

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What does MassSorter do?

MassSorter sorts, systemizes and analyzes data obtained from peptide mass fingerprinting MS-experiments where a known protein is analyzed for sequence coverage, post-translational modifications, modifications occurring during sample handling, and induced chemical modifications. MassSorter functions as a database of all the peptides detected in the experiments performed, and at the same time sorts the data according to given parameters by comparing obtained data with theoretical data. The data not recognized by this first comparison, can go through a second round of analysis where other posttranslational modifications are considered. MassSorter is meant to be a tool for small mass spectrometry laboratories that are interested in post-translational modifications of known proteins.

The basic idea is to compare experimental m/z-values from MS experiments with theoretical m/z values from a theoretical digestion of the same protein. The goal is to maximize the number of matches between the theoretical and the experimental m/z-values. MassSorter includes several tools to simplify this process:

• A theoretical protein digester that cleaves a protein sequence into peptides (both modified and unmodified versions)

• The creation of a spread sheet that summarizes all the matches and mismatches between a theoretically digested protein and data from one or several MS experiments.

• The data in the spread sheet can also be presented as an html file, where all matches and mismatches are grouped and counted according to type; unmodified match, modified match, etc. A two-dimensional model of the sequence where the detected residues are marked, is also shown.

• A tool which allows you to find unspecific cuttings: given a protein sequence and an m/z value, it finds all the possible consecutive amino acid sequences that have m/z values equal to the given search value within a selected accuracy.

• A possibility of searching for unexpected modifications by using the UniMod database (www.unimod.org).

• A tool, which given an amino acid sequence and a set of modifications, calculates the peptide mass.

• Visualization tools, including 3D modeling of the protein sequence.

• Statistical tools, which create statistics for peptide properties and for the accuracy values of an experiment.

What MassSorter is not

• MassSorter does not handle MS/MS-data.

• MassSorter does not do identification of unknown proteins by database searching using peptide mass fingerprinting (PMF). The user must use other tools to make sure the m/z-values actually come from the given protein. MassSorter builds on the assumption that the m/z-values (most likely) are from the given protein sequence.

What kind of data can MassSorter use?

MassSorter contains three ways of importing experimental data; delimited text files, XML files or by cut and paste. Delimited text files are text files where the text is ordered in columns separated by some sort of delimiter (for example empty space or ';'). XML files are more structured text files containing so-called tags explaining the content of each line. Cut and paste simply means copying the wanted date from a spreadsheet or a text file and pasting it into an experimental data table.

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What kind of data does MassSorter generate?

At present MassSorter generates three kinds of data: spread sheets, html files and text files. All the data produced in MassSorter can be saved and, at a later time, reopened in MassSorter. All information presented as spread sheets can be copied and pasted into other spread sheet programs. (But be aware that some parts of the formatting (like color-coding) might be lost in this process.) Information presented as html can be saved (or copied) and opened in other programs supporting html. The data files created by MassSorter (theoretical digestions, experimental m/z values, etc.) are saved in an internal text file format, but are easily readable and should be possible to import into other programs. (Some export formats may be supported in the future.)

To make it easy to export only the mass list of a given experiment into other programs, the list of m/z values can be transformed into a text file (see Help in the experimental data table (EDT) window).

The internal data file types are:

• *.edt - experimental data file
• *.tbt - theoretical digestion data file
• *.mmd - modification data file
• *.dst - data sheet table file
• *.enz - enzyme data file

Color Coding

The color-coding used in the DST can be altered by selecting "Edit colors" on the "View" menu. MassSorter supports two kinds of color-coding: property color and intensity grading. The default coding is property color. When comparing the m/z values, it is possible to get more than one match (within the accuracy limit) for a given experimental m/z value. The best match (smallest difference) is automatically selected as a "primary match" and the others are labeled "secondary matches". Using property color means that the primary matches have one color, the secondary matches another color and so on. This is a good way of highlighting the different types of rows in the DST. Intensity graded color-coding is based on the relative intensity of the m/z-values in the given cell. The relative intensity is divided into three classes and different colors can be assigned to each class. Intensity grading is turned on or off by selecting "Intensity grading" in the "View" menu.

It is also possible to override the color-coding and color specific rows with special colors. This can be used to highlight important parts of the DST. Simply select the wanted rows and press "Ctrl+H". A color chooser then appears. The applied colors can be reset to normal by selecting the rows and pressing "Alt+Shift+H". The highlighting can also be accessed from the "View"-menu.

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