Monday, October 16, 2017

Multiplexed plasma peptidomics!!

I stole this slide on peptidomics from this talk by Harald Tammon (see LinkedIn is good for something!)

Peptidomics is coming fo' real, yo! And this new paper in MCP jumps one of the biggest hurdles in doing these experiments! 

One of the reason peptidomics is so hard is that that processed circulating peptides are a little bit too big for metabolomics tools -- and often too small for proteomics tools. When a metabolomics person is optimizing their chromatography to separate lactic acid from alanine so they don't just shoot off the column in one single peak, that same chromatography system might not be the best for catching a singly charged peptide at 550m/z. And our tools? We rely to a huge extend on peptides accepting at least 2 charges so they provide an appropriate b/y spread for identification. +1 peptides?!? Most of the time we tell the mass spectrometer to just ignore them -- cause we aren't gonna identify them anyway.  (I wrote a post on a classic paper about this here).

How'd these authors tackle the problem? By TMT tagging everything! In general the TMT tag will add about 1 extra charge (all pH/pKa {or something} dependent) and all the sudden their normal proteomics workflow could do quantitative peptidomics!

To improve identification they used both HCD and ETHcD (forgive capitalization) and search the data with PEAKS and Byonic which are both more likely to successfully identify +1 peptides than Sequest.

How'd they do? Thousands of quantified peptides and a method that I'd follow to the letter if someone asked me to quantify changes in the global peptidome!

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