I was reading about the increase in detection of PTM phosphopeptides by the incorportaion of off-line high-pH reverse-phase fractionation and I wondered if the digestion of a proteome (not the phosphoproteome)
using pepsin + low pH conditions would increase the coverage of a proteome when used in parallel with traditional trypsin methods ?
Possible.
I read recently, that 'chymotrypsin' plus trypsin helps in better coverage of the proteome (via increased signals from proteoforms) here in MCP in 2017: http://www.mcponline.org/content/early/2017/12/08/mcp.RA117.000155.full.pdf
Thanks,
Pepsin is a random cleaver, so you end up diluting the signal by distributing it over too many random sequence lengths. Plus, you get different results with small changes in time and temperature. It's very problematic to work with. As pointed out in the above answer, mutliple digestions can improve sequence coverage. Chemotrypsin isn't the best since it has many cleavage sites. The most common pair is TRYP and AspN. Best results are to use individually and then in combination (three different digests).
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