This new study in FASEB is interesting for several reasons beyond the obvious ones like: How do you talk someone into exercising and then getting a muscle biopsy?
Doesn't ubiquitination (ubiquitylation?) just mediate protein destruction? I guess not. Geez...could any of these biological systems just be not-complicated?
For this study, Parker et al., use TMT 10-plex and 2 hour gradients on a QE Plus and Fusion system. The MS2 resolution used is 35,000 which I know some groups consider too low, but on the D30 systems like the Orbi Velos or Q Exactive Classic/Plus can make a lot more sense than getting half the number of scans because you went overkill with 70,000 depending on the experiment. In single shot and ultra-complex mixture? It can make a lot of sense.
The RAW files are all up on ProteomeXchange here.
This is a good representative image of the closest reporter ions from one of these files (keep in mind that resolution in the Orbitrap Velos is an estimation of the resolution at 400 m/z and 200 m/z in the Q Exactives, so the resolution at the tag region is much higher. Time of Flight systems, however, have uniform resolution across the m/z range, you really need a 50,000 resolution TOF to get data this good.)
Would baseline be a little better? Sure, but at the cost of 50% scans? A decision worth thinking about depending on your experiment.
This got a little off topic, but this is a cool study and a great example of doing quantitative ubiquitin signaling studies with multiplex tags!
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