Biomedical Engineering Reference
In-Depth Information
approaches based on protein quantification. This view is based
on the fact that probes for whole genomes can now be printed
on microarray chips. However, not all genes give a positive sig-
nal on the chip, presumably because not all genes are expressed
at a given time or tissue, and head to head comparison of LC-
MS-based proteomic and microarray data has shown that the two
However, although analysis of gene expression at the pro-
tein level is conceptually more powerful than is analysis of gene
expression at the mRNA level, performing LC-MS-based pro-
teomics is much more technically demanding, time consuming
and expensive than is performing microarray analyses. For these
reasons, the latter is still the method of choice for the analysis
of gene expression for most researchers. This trend may change
in the near future as mass spectrometers become less expensive,
with greater duty cycle and dynamic range, and novel quantifica-
tion methods and computer programs for their implementation
become available.
A powerful application of proteomics is in the elucidation of the
protein composition of intracellular compartments and organelles
tion aids in annotating the proteome and the genome and may
give a hint of the function of novel proteins and genes; conversely,
this is also important for understanding the function of the dif-
ferent organelles. For example, the proteome of the centrosome
has been extensively studied by quantitative LC-MS. In an influ-
ential study, Andersen and colleagues identified 23 novel proteins
resented a leap in the understanding of the composition of the
centrosome since only 60 centrosomal proteins were known prior
to this work. These findings have formed the basis for follow-
on molecular biology studies aimed at understanding the role of
A problem encountered with all MS-based organelle pro-
teomic studies is that these experiments require an initial frac-
tionation step to isolate the organelle under investigation; this is
normally carried out by differential or sucrose gradient centrifu-
gation. The sensitivity and dynamic range of modern mass spec-
trometers is such that even minor contaminants are detected in
these biochemical preparations of subcellular structures. There-
fore, a qualitative analysis by LC-MS/MS does not allow discrim-
inating true organelle components to contaminants. In order to
address this problem, researchers developed strategies to quan-
tify the enrichment of proteins in subcellular fractions relative to
the enrichment of validated markers of these fractions. This is the
2.2.2.Organelle
Proteomics
Search WWH ::
Custom Search