Tuesday, March 19, 2013

New discovery or an error?

We have seen an interesting search result that the precursor mass error plot forms two clusters. The dataset was generated with Thermo LTQ-Orbitrap instrument. Will this be related to some new science discovery or is it caused by an error? Our scientist took a closer look to find out the reason.
In the result summary view, PTM profile section, we noticed that many PSMs have the Deamidation modification.
We then look at some of those PSMs and found out that the precursor mass reported by the instrument is wrong. Instead of reporting the m/z of the monoisotopic peak, in many cases the instrument reported the m/z of the highest isotopic peak. Thus resulting a mass shift of 1Da on the precursor. Here is an example. For this charge 3 peptide, the instrument reports precursor m/z 1057.17, but the monoisotopic peak is at m/z 1056.83.
At this point, we know why there are two clusters on the plot. The software try to explain the wrong precursor mass by adding a Deamidation.

So how can we fix this? We re-run the search. This time, we checked correct precursor mass option in the data refine stage. PEAKS will try to determine the correct precursor mass by look at the survey scan instead of blindly trust values the instrument reported. Comparing the two search results, we not only removed the false Deamidation but also able to explain 10% more spectrum at 1% FDR.

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