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svkjred 09-22-2009 09:34 AM

Mass spec average peptide masses and amino acid sequence
 
Hey there

We have carried out exhaustive 2D gel electrophoresis on rice plant tissues and have identified proteins through in silico data base searches from the average peptide masses of teh mass spectrometry.

However for some protein spots the existing databases brings up no hit and these are apparently the most responsive proteins and look very novel.

Now i have the average pepetide masses of these proteins and looking into Expasy (both Profound and Mascot) gives me just a couple of peptide hits with very low significance probability.

Is there any way that the amino acid sequence from the average pepetide masses be deduced something like using tripsin restriction sites and the molecular masses entangling the possible amino acid sequence or is it just impossible.

Thanks a lot
Krishna
Post doc
International Rice Research institute

danfive 09-22-2009 05:29 PM

Re: Mass spec average peptide masses and amino acid sequence
 
If you have the m/z values for the peptides of interest, you should compile them into a list. Then run (or contract) an MS/MS run--the first MS measures the m/z value and then sends that peptide into a collision chamber--breaking it into its individual amino acids--then into the second MS that tells you the amino acid makeup and sequence of your unknown peptide.

This is the great benefit of MS/MS it sequences the unknown peptides from just MS.
Prior to this run you do trypsinize the "spot", reduce and alkylate then run it.


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