Biomedical Engineering Reference
In-Depth Information
contribute to the accumulation of protein aggregates during dis-
eases (
39
). The stable properties of carbonylated proteins make
them interesting analytical targets and they have been used as
markers of oxidative stress (see Chapter 1). Carbonylation is a
consequence of a reaction between reactive oxygen species (ROS)
and proteins and it keeps a record of bursts of oxidative stress in
a specific cellular location, whereas ROS itself and downstream
products like H
2
O
2
are quickly metabolized. Measurement of car-
bonylation can be performed at different levels of sample com-
plexity starting with a crude mixture of proteins. Overall
measurement of the extent of protein carbonylation can be assayed
on Western blots (see Chapter 17). Using MS, the carbonylated
proteins can principally be identified directly in a liquid mixture.
However, it might prove difficult since there are many combinations
of carbonylation products, each at a relatively low level. Therefore,
a common strategy is to increase the signal of carbonylated proteins
by first enriching for the carbonylated proteins, which in some
cases even enables identification of the amino acid site of carbony-
lation (
40
). Enrichment using affinity chromatography has been
applied using different derivatization agents: DOTA (
41
), Girard
P reagent (
42
) and biotin hydrazine (
40, 43
).
Large-scale protein carbonylation studies have been applied
to, for example, human post mortem brain samples of
Huntington disease (
44
) and on the mouse model PS1+AbPP
of Alzheimer's disease (
45
), and have thus contributed to the
understanding of neurogenerative diseases.
5. Metabolomics
METABOLOMICS, in some scientific circles also called
METABONOMICS, is a growing OMICS branch, which has
the aim to quantitate - as many as possible - metabolites from
body fluids or cell extracts and thus describe the status of cells
or organisms with respect to metabolic activity. Currently, the
major employed methodologies are MS and nuclear magnetic
resonance (NMR) spectroscopy. MS is able to detect a larger
number of different metabolites, but laborious upstream frac-
tionation strategies must be used focusing on fractions of metab-
olites with similar gross chemical properties (like hydrophobicity)
and then combining the data to a more comprehensive picture.
In contrast, NMR does not require upstream fractionation, but
it covers clearly fewer metabolites, is less sensitive and identifica-
tion is more challenging. METABOLOMICS is usually dealing
with a limited fraction of the many metabolites present in bio-
logical systems, but the big step from analyzing one or a few
metabolites to a more broad selection has definitely been made
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