Xque - 9-8-2006 at 01:34
I'm starting to understand the methods of protein analysis, when one thing came to my mind...In determining the sequence of a protein there's two ways
to go (AFAIK 
), you can either cleave the protein into smaller peptides, then
sequence them using the Edman degradation and then connect the peptides into the proteins sequence using overlap peptides, obtained by a second
cleavage with another enzyme. Or you could cleave the proteine, determine the masses of the peptides by MALDI-TOF and then look up the peak list in a
protein database.
Have I understood these concepts correctly? If yes, how do you decide which one to use for your analysis? What are the ups and downs of each of these
methods? Are there other methods you could use for determining the sequence of a protein?
I'm sorry for all the questions, but I'm really getting into the analytical biochemistry 
[Edited on 9-8-2006 by Xque]
silonyl - 30-8-2006 at 06:59
Your analysis of the underlying concepts is quite accurate. Now it's been a while since I've been exposed to hands-on analytical biochem, but my
understanding of the differences between these two methods is as follows:
Analysis by MALDI-TOF MS requires minimal hands-on laboratory work, making it highly attractive for well-funded and high-throughput applications.
Although it requires only minimal sample preparation - I think cleavage may even occur during the desorption/ionization process, making that step
unnecessary - the actual MALDI MS is a very expensive instrument and requires dedicated trained personnel to maintain and operate it... so this
technique is really only available in-house to large corporations and institutions.
The Edman Degradation method requires less in the way of instrumentation - only chemical facilities and the ability to perform chromatography or
electrophoresis - but it requires a LOT more investment of human time. My guess is that this method is more popular at smaller companies and
institutions where tech/student labor is cheap and abundant but the funds to create a high-tech instrumentation facility are not.
Hope this helps.