"Frontmatter". In: Plant Genomics and Proteomics

P
ROTEIN
C
HARACTERIZATION
In many ways the approach to generating and understanding the protein
sequence has come full circle. Originally it was thought that the way to iden-
tify the sequences of genes would be by sequencing the proteins, and Sanger
was awarded the Nobel Prize (1958) for devising methods of getting peptide
3 8
2. T
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B
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O O L B O X
— A
C Q U I R I N G
F
U N C T I O N A L
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E N O M I C
D
ATA

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sequences. However, it then turned out that DNA sequencing was much
faster, leading to the explosion of DNA sequence data (and another Nobel
Prize for Sanger in 1980), with the protein data coming from the nucleic acid
sequence (usually from the cDNA sequence because, even now, gene pre-
diction programs still need much development to be applied with confi-
dence). The application of mass spectrometry to protein characterization and
identification has improved the amount of information that can be obtained
from biological samples, including the ability to do peptide sequencing on
very small samples. The proteome can be described as the full protein com-
plement of an organism, cell type, or tissue at any one moment in time. Pro-
teomic studies are still in their infancy but are becoming more important and
pervasive. One key technology for proteome analysis is two-dimensional gel
electrophoresis (2-DE), which has the capability to separate very complex
protein mixtures. These mixtures can contain thousands of components with
different physicochemical properties and abundances. A second key tech-
nology is mass spectrometry, which is useful for proteome analysis because
it has a sensitive detection range and can be used for high-throughput iden-
tification. The results obtained from these techniques can be applied to gene
and protein database searches and can also be used to identify posttransla-
tional modifications of proteins (information that is impossible to acquire
from nucleic acid studies). The most important ionization techniques for pro-
teomics, matrix-assisted laser desorption/ionization (MALDI) mass spec-
trometry (MS), and electrospray ionization (ESI), have been continuously
improved so that MS plus MS/MS data have enhanced the protein identifi-
cation capabilities of the methods (Figure 2.7) (Pandey and Mann, 2000).
Proteome analysis is now a complementary and potentially coupled
technology to transcription profiling. However, proteome-based studies
must be carefully designed and performed to ensure reproducible analysis.
In particular, the selection of the tissue and its preparation are crucial steps
in proteome analysis.