Initial proteome analysis of mature barley seeds and malt

Abstract

Several barley (Hordeum vulgare) cultivars are used in the production of malt for brewing. The malt quality depends on the cultivar, its growth and storage conditions, and the industrial process. To enhance studies on malt quality, we embarked on a proteome analysis approach for barley seeds and malt. The proteome analysis includes two-dimensional (2-D) gel electrophoresis, mass spectrometry, and bioinformatics for identification of selected proteins. This project initially focused on proteins in major spots in the neutral isoelectric point range (pI 4-7) including selected spots that differ between four barley cultivars. The excellent malting barley cultivar Barke was used as reference. Cultivar differences in the 2-D gel spot patterns are observed both at the seed and the malt level. In seed extracts one of the proteins causing variations has been identified as an alpha-amylase/trypsin inhibitor. In malt extracts multiple forms of the alpha-amylase isozyme 2 have been identified in varying cultivar characteristic spot patterns. The present identification of proteins in major spots from 2-D gels includes 27 different proteins from 42 spots from mature seed extract, while only three specific proteins were identified by analysing 13 different spots from the corresponding malt extract. It is suggested that post-translational processing causes the same protein to occur in different spots.

Complexity of the human whole saliva proteome

“…In this frame, nearly 30 spots might be associated to unglycosylated forms, and at least 20 spots to glycosylated forms. Thus, α-amylase appears to display a diversity which is far larger than previously reported (7,20,21 (18). For most α-amylase spots that are observed at various MW lower than expected, other mechanisms must be taken into account.…”

Developmental Analysis of Maize Endosperm Proteome Suggests a Pivotal Role for Pyruvate Orthophosphate Dikinase

“…About 200 protein spots were identified in successive studies, belonging to primary metabolism, cell rescue and defense, and protein folding. Specific accumulation patterns were observed during grain development and germination (Finnie et al, 2002;Ostergaard et al, 2002Ostergaard et al, , 2004. A recent study in wheat focused on albumins and globulins, excluding the abundant storage proteins glutenins and gliadins; 254 proteins were identified and their accumulation profiles were compared at 10 and 36 d post anthesis.…”

Integration of the barley genetic and seed proteome maps for chromosome 1H, 2H, 3H, 5H and 7H

“…Proteins in 100 or 200 μL extract (100 or 200 μg protein) for pI 4-7 or 6-11 gels, respectively, were precipitated with 4 volumes of acetone at −20°C for at least 24 h. Precipitated proteins were redissolved in the appropriate IPG rehydration buffer prior to the first dimension, as previously described (Østergaard et al 2002;Bak-Jensen et al 2004). For separation of proteins in the pI ranges 4-7 and 6-11, immobilised pH gradient 18 cm IPG strips were run on an IPGphor unit or a Multiphor II, respectively (GE-Healthcare), and the second dimension SDS-PAGE gels (12-14%, 18×24 cm, GE-Healthcare) were run on a Multiphor II as previously described (Østergaard et al 2002;Bak-Jensen et al 2004). Gels were stained with silver nitrate (Heukeshoven and Dernick 1985).…”

“…Proteins in 26 of the spots were already identified as part of ongoing barley seed proteome analyses (Bak-Jensen et al 2004Finnie et al 2002Finnie et al , 2004Finnie et al , 2006Finnie and Svensson 2003;Laugesen et al 2007;Østergaard et al 2002. Proteins in an additional 22 varying spots were successfully identified by mass spectrometry (Supplementary Table S2) resulting in the characterisation of 48 out of the 69 variable spots.…”