Liquid chromatogram coupled with mass spectrometry (LC-MS) is a pre-dominant platform for proteomics research. Top-down proteomics using LC-MS has been increasingly applied for analyzing intact proteins to study genetic variation, alternative splicing, and post-translational modifications (PTMs) of the proteins, where each intact protein may have various distinct forms of variation or modification termed as proteoforms. However, only a few tools have been developed to perform charge state deconvolution, average molecular weight determination and quantitation of intact proteins from LC-MS spectra. Though Decon2LS and MASH Suite Pro have been available to provide intra-spectrum mass deconvolution and quantitation analysis, manual processing is still required to quantify proteoforms based on abundances obtained from multiple spectra. Currently there are few automated tools for large-scale proteoform quantitation based on inter-spectrum abundance in top-down proteomics. In this study, we present the first fully automated tool, called iTop-Q (intelligent Top-down proteomics Quantitation), for top-down proteomics quantitation using XIC abundances. Instead of utilizing single spectrum for proteoform quantitation, iTop-Q constructs extracted ion chromatograms (XICs) of possible proteoform peaks across adjacent MS1 spectra to calculate abundances for accurate quantitation. Notably, iTop-Q is implemented with a newly proposed algorithm, called DYAMOND, using dynamic programming for charge state deconvolution, instead of using THRASH as in other tools. Furthermore, iTop-Q performs proteoform alignment to support quantitation analysis across replicates/samples. The performance evaluation on an in-house standard data set and a public large-scale yeast lysate data set shows that iTop-Q achieves highly accurate quantitation, more consistent quantitation than using intra-spectrum quantitation. Furthermore, the DYAMOND algorithm is suitable for high charge state deconvolution and can distinguish shared peaks in co-eluting proteoforms.
- The User Guide of iTop-Q (pdf)
- For users using Windows Vista or Windows 7, please make sure that the "Privilege Level" of Compatibility has been checked.
(Right click on the icon of iMet-Q->Compatibility->Privilege Level: Run this program as an administrator)
|The recommended system environment is:
- Operating System: Windows XP, Windows 7, 8, 10 and Windows Server 2008, 2012
- CPU: Intel® CPU 2.40 GHz or faster
- Hard Disk: depending on the input file size. It would be safe to have the free disk space at least a few times the total size of input files.
- Memory: depending on the input file size. As a rule of thumb, 4GB or more is suggested. Larger input files will be more efficiently processed with larger free memory space.
- Microsoft .Net Framework: .Net 4.0 or higher
- Screen Resolution: 1024*768 or higher
If you are unable to open the tutorial video, you may also download it (link).
||V1.000 is available
- iTop-Q is officially on-line.