CH908, Problem set 8. Collisionally Activated Dissociation (CAD) of peptides 1. Memorize the structures of the 20 naturally occurring amino acids. 2. In the mobile proton model of peptide fragmentation, the initiating event (after collisional activation of the molecule) involves a proton transfer. From where? To Where? The proton is normally on Arginine > lysine > histidine > n-terminus > backbone amides. If it's on arginine or histidine, it's stuck and doesn't move, so fragmentation of such peptide ions is poor. If it's elsewhere, it can transfer to the backbone amide to initiate cleavage. This results in a one Da transfer from nominal "b" ions to nominal "y" ions. 3. Calculate the CAD fragments that would be expected from the following peptide sequence: RGHTTPSDELWIVK. What can you say about expected relative cleavage intensity? Create a table of the expected fragments and mark them with "strong, medium, weak" for expected intensities. Fragmentation next to proline (b6, y9) will probably be intense, as will cleavage next to aspartic acid (b9, y6). For the rest, it's safest to just call them "medium", except for cleavage at glycine which will be weak. 4. Calculate the CAD fragments that would be expected from the following peptide sequence: RGHTpTPpSDELpYIVK. Note: pS, pT, and pY refer to phosphoserine, phosphothreonine, and phosphotyrosine, respectively. What can you say about expected relative cleavage intensity? Create a table of the expected fragments and mark them with "strong, medium, weak" for expected intensities. Since this peptide is triply phosphorylated, it's likely to be poorly observed in positive ion mode, but nicely detectable in negative ion mode. In positive ion mode, the same rules as above apply, but you should also see several intense losses of phosphoric acid (-98 Da), several water losses. Because of these losses, other backbone fragmentation will probably be weak, but it will be evenly distributed because of the abundance of mobile protons. 5. What's wrong with this picture? The z and x ion cleavages are wrong, they are swapped. The rest are correct. a/x, b/y, c/z go together.