Mass Spectrometry in Systems Biology - Systems Biology: Networks, Models, and Applications - page 19

an m / z value of a precursor ion of low abundance [62]. Alternatively,
MALDI-MS/MS could identify the presence of a neutral loss (for
example, neutral loss of H 3 PO 4 from Ser/Thr phosphorylated pep-
tides). Then, one could employ targeted MS 3 for structural elucidation
of the peptides showing the neutral loss [63]. Correspondingly, in an
LC-MS/MS arrangement, data-dependent neutral loss on an ESI linear
ion trap performs the same experiment for hypothesis-free analysis.
Either evaluation of differential quantities of peptides, MS/MS spectra,
or neutral loss scans can be used as criteria for hypothesis-driven mass
spectrometry. Graber et al. [64] proposed an integrated workflow based
on the use of MALDI to make decisions on follow-up experiments
using ESI. The workflow relies on a relational database that organizes
the acquired data for further result-driven mass spectrometry.
Recently, Venable et al. [59] developed a data-independent strategy
for the acquisition of tandem mass spectrometry data for large-scale
quantitative proteome analysis using fast scanning instruments like the
linear ion trap. Data-independent tandem MS method indiscriminately
collects MS/MS spectra without MS survey of the data as required by
data-dependent acquisition.
Ultimately, the basic function of any proteomic platform (hyphothe-
sis-driven or hypothesis free) is elucidation of structures of peptides as
well as precise amino acid localization of posttranslational modifica-
tions as a function of cellular events. From these data, we can then
reassemble the scaffold of proteins by describing their sequence cover-
age, modifications, and relative expression level, parameters that will
eventually describe the biological system.
Table 1.1 summarizes the different proteomics platforms, separation
techniques, and MS instrumentation used in papers cited in this chapter.
The first three sections of the table summarize analytical techniques
used for analysis of proteome and large-scale identifications of modi-
fied peptides (phosphorylated and/or other modifications). Analysis
of modified peptides requires in some cases preparative protocols of
protein biochemistry (column I). After proteolytic digestion of proteins,
the peptide mixture is further simplified by preparative offline chro-
matography for peptide quantification or identification of modifications
(column II). Column III summarizes the combinations of LC-MS used.
For proteome analysis, MudPIT presents widespread applicability with
no other sample preparation other than digestion of lysates or cellular
organelles. In order for single LC coupled with MS/MS to analyze
samples of the same complexity, one needs to rely either on extensive
sample preparation (column I or II) and/or employ MS instruments
with increased mass accuracy and resolving power (Q-TOF, FTICR).
On the other hand, column IV emphasizes the role of ion trap MS as the
workhorse of proteomics studies, particularly its scanning capabilities
for large-scale analysis of peptides. Analysis of modified peptides