Sunday, March 30, 2014
Byonic in Proteome Discoverer
As I mentioned earlier, one of my favorite MS/MS programs ever, Byonic, has been seemlessly integrated into the Proteome Discoverer 1.4 environment. If you haven't heard of Byonic, I wrote a couple posts on it over the last 2 years. Essentially, Byonic gives us the opportunity to study PTMs well, thoroughly, and confidently. And you can just tack it into PD now.
Specific focuses are available for glycopeptides! What else does glycopeptide analysis? Sure, there are some packages out there, but my dog, this is EASY. I have some glycopeptide files here now that I may process and update you on, but my sample queue is loaded for at least the next 3 days. The sounds of the CPU fans in this room, lol!
Wildcard search! Fully customizable wild card search! Wooooohoooo! Okay, so you are thinking "big deal, right? We can wildcard search in Mascot and some other software." Sure. We. Can. And not to poo on anybody, but this works. And it works really really well.
Proof? You know those crosslinkers that were developed at the University of Victoria Protein Centre? (Described in this paper). You should. Cause they're awesome. Isotopically labeled and CID cleavable. The idea is that when two proteins are in association, you crosslink them with this compound. The crosslinks come in two types, the heavy and the light, but otherwise completely identical. They are exactly 8.00 (I forget) Da apart. You can then use the Mass tags function on the Orbitraps to, basically, do a gas phase enrichment for the low abundance crosslinked counterparts. Otherwise, you basically can't see them.
So, I took a file that was treated this way and ran it. Consider this. Its an extremely low abundance mod. Oh, and its >500 Da in size. What are the chances that Byonic could find a mod like this? So, I gave Byonic this file and didn't tell it about the mod. At all. I just said "do a wildcard search up to 600 Da". It took a while. Imagine that it had to look at every amino acid possibility and up to 600 Da heavier! What are the combination? It took a few hours.
But, you know what it found?
The crosslinked peptides. Exactly where they should be. Exactly what they should be.
Unimpressed? Try this, or something like it with another program. Let it search its maximum allowable "wildcard" type mass window. Then email me if it you don't have any false hits, because by doing a search like this you have inherently dumped a whole ton of chaos into your search and you have made your FDR engine do impossible numbers of calculations. I've never seen a search like this come back without silliness in it. There are too many variables. And Byonic just does it. Correctly. Oh, and it automatically (unless you turn it off) throws in a target decoy search as well as searches common contaminants (it appears to be the cRAP database).
This little company in San Carlos is quietly writing the best shotgun proteomics software I have ever seen. And if you are using PD, you already know how to use it.
TL/DR: Byonic is awesome. I have no idea how they do it, but it is awesome.