MS/MS and CID MS/MS
One of the most powerful tools available to the mass spectrometrist is MS/MS.
Whereas fragmentation is virtually absent in electrospray mass spectrometry, it can
be deliberately induced by MS/MS techniques. Tandem MS employs collision-
induced dissociation (CID) to fragment a precursor ion. For a sample mixture (even
after chromatography, a mixture of different species can be present), the first
quadrupole mass analyzer (Q1 in the QSTART XL) generates a spectrum of ions.
The ion of interest is then selectively allowed to progress into the collision cell (Q2),
containing an inert gas typically argon, which is allowed to collide with the selected
ion. This induces fragmentation, caused by collision-induced dissociation (CID) and
the resulting ions are then analyzed as product ions. Sequence as well as other
structural information can be derived from these fragments. Tandem MS/MS can be
used to elucidate the structure of a variety of small to medium sized molecules,
including organic molecules, lipids and fatty acids, peptides, carbohydrates,
oligonucleotides, as well as DNA and RNA adducts, etc.
In the past a few years, researchers have begun to use mass spectrometry to
determine peptide sequence. Improvements in machinery sensitivity as well as
software now allows for the confirmation of peptide sequence with virtual certainty.
When fragmented, a series of ions are produced, termed a, b, and c ions for those
with N-terminal charge and termed x, y and z ions for those fragments with C-
terminal charge. The b and y ions are the ions generated when the fragments are
due to the cleavage at the peptide bond, whereas the a, c, x, and z ions are
generated when cleavages occur on the N-terminal side of the peptide bond carbon
(a and x) C-terminal side of the peptide bond amine (c and z). These ions are then
matched up with known amino acids and amino acid fragments, and the mass
difference between consecutive ions reveals the identities of the consecutive amino
acids. From this map, an accurate peptide sequence can be determined.
The QSTART XL is so accurate in determining mass of the various CID fragments,
that with the exception of leucine and isoleucine, two amino acids with identical
molecular weights, all other amino acids can be differentiated. Even glutamine, with
a mass of 128.0586 and lysine, with a mass of 128.0950, can be differentiated.
Similarly, various amino acid modifications can be determined due to the gentle
nature of ESI (versus MALDI, which tends to remove some modifications, such as
phosphates). These include phosphorylation, glycosylation, acylation, nitronation,
and many others.
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