Tuesday, October 14, 2014
Global analysis of protein structural changes!
This one is really smart. And I both expect and hope to give this a try soon.
Proteins have 3-dimensional structures. What the protein looks like in 3D is critical to what the protein does, who it interacts with, and all sorts of other stuff (yes, there is better terminology secondary through tertiary...I took biochem...a long time ago...)
By necessity (or so we thought) shotgun proteomics has always ignored this fact. We denature the proteins down to their 2 dimensional straight chains and then digest them from there. As a consequence we only get part of the story, maybe a small part of the story.
This new paper in Nature Biotechnology suggests that we can go after this information, and we don't have to buy a fancy NMR or tons of crazy reagents to do it -- chances are, we have everything we need to find out (in some sense, at least) how our protein 3D structures are changing from one sample to another.
Check out this example I found on Google Images (I couldn't find the original author. If this is you, I apologize and leave a comment to get credit for this). In this awesomely simple image this protein has 2 states, Open and Closed. In the Open state, if we look right where the arrow is pointing we have access to this nice middle region. If we were to digest this protein in its native state, we would be able to see this middle region. Skip to the right. Now the protein sequence is closed. The amino acids in the middle are protected by the rest of the protein chain. My enzyme of choice can't access this site in the Native Closed protein and no peptides from that area would show up in my global proteomic analysis.
Simple and elegant, right?!?!?! So, what we need to do is carefully extract our proteins in their Native state in condition 1 and in condition 2 and digest them softly. The authors of this great paper then denature everything and digest under normal conditions. By using 2 different enzymes, they can find that exposed regions in the 2 conditions disappear (or are just cut much smaller!) and the regions that don't change between the 2 conditions appear the same between the 2 samples.
Papers like this are the reason I get out of bed...that and the Pug threatening to pee on my floor....
Big thanks go to Dr. Sreelakshmi for tipping me off to this paper!
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