news
ANALYTICAL CURRENTS
Isolating glycoproteins with lectins John Wilkins and colleagues at
other well, allowing identification Unassigned
the Manitoba Centre for Proteomics, the University of Manito-
of 132 unique membrane glycoproteins. Only 26 proteins were
Mitochondria
ba, and Beavis Informatics have developed a method for isolating
captured by both columns. Analysis of the eluted proteins from the
Plasma membrane
membrane glycoproteins using lectin affinity chromatography. Al-
columns showed a 150–200-fold Lyso./Golgi/vesicle
though membrane proteins are
enrichment for glycosylated proteins from the whole-cell lysates.
Endop. retic.
thought to comprise almost a
The entire set of isolated proteins 0
third of the eukaryotic genome, the present state of knowledge of these proteins is limited because
10
20
30
40
50
Distribution of membrane proteins identified (%)
Distribution of membrane glycoproteins purified by lectin affinity chromatography in various cellular compartments.
of the difficulty in isolating them. Membrane proteins are often modified
covered a wide range of molecular weights and isoelectric points. Wilkins and colleagues analyzed the cellular distribution of
lectins recognize distinctly different sugars.
the membrane glycoproteins and found them
Whole-cell lysates were loaded on ei-
present in the plasma membrane, Golgi vesi-
with large chains of sugars to produce glycoproteins. In their approach, Wilkins and
ther Con A or WGA columns. The bound
cles, endoplasmic reticulum, and mitochon-
colleagues used lectins, proteins that spe-
proteins were eluted and subjected to pro-
dria. Thus the investigators concluded that
cifically bind to sugars, to capture mem-
tease digestion. The digested peptides
their approach with lectin affinity chroma-
brane glycoproteins. The investigators
were separated by HPLC and identified
tography could be very useful for the pro-
used two lectins, concanavalin A (Con A)
by MALDI MS/MS.
teomic analysis of this essential class of
Wilkins and colleagues found that the
and wheat germ agglutinin (WGA), as matrixes for affinity chromatography. The two
two lectin matrixes complemented each
proteins. (J. Proteome Res. 2004, 3, doi 10.1021/pr049937f)
Dioxins in fish Marc Bassompierre and colleagues at the Royal Veterinary and Agricultural University in Denmark have developed a rapid and inexpensive way to screen for dioxins in fish. The method, which relies on IR spectroscopy and multivariate analyses of fatty acid patterns, could be used for routine monitoring of dioxins in fish and fish products. Recent food safety regulations in the European Union require increased monitoring of dioxins in food, particularly in animal and fish products. The levels of dioxins and furans in food are typically determined by highresolution (HR) GC/GC/MS, which is too expensive for routine monitoring 256 A
and quality-control applications. Dioxins bioaccumulate in fat, and lipid profiles have been shown to reflect dietary patterns. Now, it appears that fatty acid profiles can also be used as a natural tracer for dioxins. Bassompierre and colleagues measured dioxins in 64 fish meal samples from Northern Europe using HRGC/ GC/MS. The dioxins were expressed as the sum of the toxic equivalencies (TEQs) of 2,3,7,8-TCDD, the most toxic dioxin congener, in accordance with World Health Organization (WHO) guidelines. The level of dioxins in the fish meal samples ranged from 1.1 to 47.1 ng TEQ-WHO/kg fat, with an
A N A LY T I C A L C H E M I S T R Y / A U G U S T 1 , 2 0 0 4
average of 15.3 ng TEQ-WHO/kg fat. The fat content and fatty acid profiles of each sample were measured using nearIR spectroscopy and GC, respectively. Using a multivariate model with three partial least-squares regression components, the researchers discovered that 13 fatty acids, ranging from C14:1 n-5 to C24:1 n-9, in fish were positively correlated with the level of dioxins. The approach is about 1000 faster than conventional analyses for dioxins, and it requires only chemometric software and fatty acid measurements, which are typically performed using GC with flame-ionization detection. (Analyst 2004, 129, 553–558)
