Biology Reference
In-Depth Information
It is amazing how much information one can get by merely “weighing
things.” However, when one realizes that MS offers enough resolution to
distinguish a mass difference in the range of a single proton to half a mil-
lidalton using Fourier transform ion cyclotron mass spectrometry (FT-
ICR-MS) (Marshall et al ., 1998), then it is not so surprising why this
technique is so powerful.
Most of the variations we try to track as indicators of abnormal phys-
iology can be related to a mass change. When two molecules interact or
are modified, there is a corresponding change in mass (except rearrange-
ment reactions). When peptides or proteins are modified chemically,
there would also be a change in mass seen as an increase or decrease
attributed to the moiety added or removed, respectively. Because MS
allows directed fragmentation, even isomers can be distinguished based
on the different fragments generated as a function of structure; hence,
products of different masses are obtained giving insight into chemical
structure.
Analytes or samples to be studied by MS can be solid, liquid or gas as
starting materials as long as charged analytes can be generated from them,
because MS depends on gas-phase mass separation and detection of ions.
Early methods of ionization were limited to analysis of small molecules.
With the introduction of fast atom bombardment (FAB) (Morris et al .,
1981), it became possible to ionize intact peptides. However, it was not
until soft-ionization methods, namely, ESI (Fenn et al ., 1989, 1990) and
MALDI (Karas et al ., 1988; Chaurand et al ., 1999) were developed, that
rapid and facile applications to large biomolecules were made routinely
possible.
Peptide mass fingerprinting
MALDI coupled to the time-of-flight (ToF) analyzer has been a workhorse
for the strategy called peptide mass fingerprinting (PMF). This is based on
the fact that proteins having unique sequences have distinct patterns of
cleavage sites for a given enzyme. The cleaved protein therefore yields
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