Biology Reference
In-Depth Information
ligand libraries (CPLL) (ProteoMiner as well as a homemade
library of reduced polydispersity) at three different pH values. The
CPLL technique offers a unique increment in sensitivity for low
abundance protein species. Therefore, they identifi ed 20 and 40
unique gene products from barley and
Saccharomyces cerevisiae,
respectively, by the mass spectrometry analysis of the recovered
fractions. Avenin-like protein-a was identifi ed in beer and the pro-
tein enriched using silica gel from beer foam by nano-ESI-MS/
MS analyses [
11
,
12
,
20
]. Weber et al. [
20
] analyzed eight differ-
ent beer products by LC-MS/MS after digestion with trypsin and
identifi ed B-, D-, and
-hordeins and several albumins such as pro-
tein Z and LTP1. Colgrave et al. [
12
] analyzed wort, beer, and
hordeins extracted from barley fl our by LC-MS/MS and identifi ed
B-, C-, D-, and
γ
-hordeins and avenin-like protein-a. These results
suggested the feasibility of alternative techniques to determine
allergen and gluten markers in beer instead of ELISA-based meth-
odologies. Table
1
summarizes a list of proteins identifi ed in beer.
γ
2DE has been used to analyze barley grain and/or malt proteins
for over 20 years and mass spectrometry has only recently been
applied. Beer proteins consist predominantly of barley albumins
and globulins, which are released into wort, and subsequently,
appear in small amounts after processing (20-600 mg/L) [
21
].
Only approximately one-third of the total protein content remains
in beer after degradation by mashing and precipitation during wort
boiling. Therefore, quantitative and qualitative changes of proteins
occur in malting and brewing processes. These changes are also
signifi cantly caused by malt modifi cations. Silva et al. [
22
] investi-
gated the protein fractions of malt, wort, and beer from two barley
cultivars by SDS-PAGE and reversed-phase HPLC (RP-HPLC),
and showed that protein profi les from the wort samples between
the two cultivars tested were different; however, those between
beer samples were similar. Perrocheau et al. [
8
] analyzed barley
grain, malt, and beer proteins by 2DE and mass spectrometry to
investigate proteome changes during malting and brewing. In
these processes, most of the heat-stable proteins are disulfi de-rich.
A comprehensive proteome map constructed for sweet wort by
Iimure et al. [
23
] consisted of 63 identifi ed proteins out of 202
protein spots on 2DE images, which were categorized into 20 pro-
tein species. Colgrave et al. [
12
] identifi ed 27 proteins including
hordeins in the wort sample by LC-MS/MS analysis.
2.2.3 Proteome Analysis
of Wort
Barley proteins are digested in malting and mashing processes by malt
proteases, and subsequently amino acids and low molecular weight
polypeptides are present in the wort. 2DE cannot be used to analyze
low molecular weight polypeptides, i.e., below approximately 8 kDa.
To detect low molecular weight polypeptides, we should use alterna-
tive methods such as LC-MS/MS in addition to 2DE analysis.
2.3 Analysis of Low
Molecular Weight
Polypeptides in Beer
and Wort
