Separation and characterization of basic barley seed proteins

Abstract

Basic proteins in barley starchy endosperm from developing seeds were separated by two-dimensional (2-D) nonequilibrium pH gel electrophoresis. Total as well as partial extracts were analyzed. Edman degradation sequencing and immunological detection were performed after transfer of separated proteins onto membranes. Only one protein could be analyzed by N-terminal sequencing of blotted and separated proteins from the total extract. Fractionation of extracts was done using cation exchange chromatography, concanavalin A and heparin affinity chromatography. Internal sequences were determined after in-gel cleavage of proteins using trypsin or cyanogen bromide and separation of the fragments by reversed-phase chromatography or in a gel electrophoresis system for peptide separation. This resulted in a new protocol for obtaining internal sequences from proteins separated by 2-D electrophoresis. A total of 16 sequences, including nine internal sequences, were analyzed, permitting the identification of ten proteins, including five that appeared to have a blocked N-terminus. An additional protein was identified using immunological detection. Three protein sequences remained unidentified. Separated proteins were also analyzed with a glycan detection method.

What is in a Beer? Proteomic Characterization and Relative Quantification of Hordein (Gluten) in Beer

“…Among the most abundant proteins detected were the previously reported B3-hordein (), γ-3-hordein () and the predicted γ-1-hordein (). Likewise, the D-hordein () was detected in abundance.…”

Plant proteome analysis

“…Identification of selected spots revealed proteins already known to be associated with drought stress, but also others were identified such as caffeate O-methyltransferase detected only in stressed plants indicating increased lignification in response to the stress treatment [25]. These and many other studies have demonstrated the capacity of 2-DE to document genetic variability and to distinguish between lines and varieties, e.g., when analyzing barley seed and malt [26,27] or wheat grains [28]. Positional shifts of proteins were observed in the 2-D gel analysis of segregating families of maize, barley, pea, and maritime pine [29].…”

Probing heat-stable water-soluble proteins from barley to malt and beer

“…In contrast with the plant model for dicots and monocots, i.e., Arabidopsis thaliana and Oryza sativa, respectively, the lack of complete genomic databanks for barley limits the efficiency of proteomics for this application. However, preliminary maps of soluble barley seed proteins, i.e., albumins and globulins, have been obtained either on mature [5,6] or developing and germinating grains [7][8][9]. Recently, more specific protein maps have been obtained on the major compartments of the seed, i.e., starchy endosperm, aleurone layer, and embryo [10].…”