Protein Disulfide Isomerase (PDI1-1) differential expression and modification in Mexican malting barley cultivars

Abstract

Barley malting quality depends on seed characteristics achieved during grain development and germination. One important parameter is protein accumulation in the mature seed, which may vary between cultivars. Here we conducted a protein pattern analysis in the range of pI 4–7 of mature grains from five Mexican barley cultivars, commonly used for malt and beer production. Reproducibly distinct protein spots, separated by 2D SDS PAGE, were identified by mass spectrometry and considered as potential markers for cultivars with distinct seed protein accumulation. The expression patterns of glutamate decarboxylase (GAD) and protein disulfide isomerase (PDI1-1) were followed at transcript level during grain development for three independent growth cycles to establish whether differences between cultivars were reproducible. Quantitative determination of PDI1-1 protein levels by ELISA confirmed a reproducibly, distinctive accumulation and post-translational modifications between cultivars, which were independent of plant growth regimes. According to its impact on differential storage protein accumulation, we propose the PDI1-1 protein as potential biomarker for Mexican malting barley cultivars.

Identification of differential protein recognition pattern between Naegleria fowleri and Naegleria lovaniensis

“…All reagents were purchased from Sigma‐Aldrich Química, Mexico, unless otherwise stated. Protein extraction was done according to previous reports by Hajduch et al and Herrera‐Diaz et al 33,34 with some modifications. Briefly, the amoebas ( N fowler i and N lovaniensis ) were centrifuged at 4000 g for 6 minutes, supernatants decanted and the resulting pellet was suspended in 10 mL of buffer containing 50% [v/v] of phenol pH 8.8, and 50% of 0.1 mol/L Tris‐HCl pH 8.8, 0.9 mol/L sucrose, 0.01 mol/L ethylene diamine tetra‐acetic acid (EDTA), 0.4% [v/v] 2‐betamercaptoethanol and EDTA‐free protease inhibitors (CompleteTM; Roche Molecular Diagnostics).…”

“…Previously obtained proteins were separated by two‐dimensional electrophoresis according to the method described by Herrera‐Diaz et al 34 with some modifications. Briefly, 500 µg of total protein extracted from N fowleri or N lovaniensis was applied on a 11 cm polyacrylamide gel strip with a pH immobilizer gradient (IPG BIORAD) with pH range of 3‐10 or 7‐10, respectively, for 10 minutes, at room temperature, in a rehydration tray (Immobiline DryStrip Reswelling Tray, BIORAD).…”

Proteomic profile of tepary bean seed storage proteins in germination with low water potential

“…To demonstrate the quality of the 2-DE analysis and obtain a large number of spots, 1 g of cotyledons from seed cotyledons germinated at 72 h water (control) and − 0.49 MPa were placed in 6 mL of phenol pH 8.8 and 5 mL of extraction buffer (100 mM Tris HCl, pH 8.8, 10 mM EDTA, 900 mM sucrose and 0.4% 2-betamercaptoethanol). Total protein was extracted and separated by two-dimensional electrophoresis following the methods previously described by [ 25 ] with slight modifications.…”

Endomembrane‐mediated storage protein trafficking in plants: Golgi‐dependent or Golgi‐independent?

“…Surprisingly, PDI1-1 loss improves rice flour characteristics and food processing properties [82]. In barley, PDI1-1 has also been suggested as a candidate biomarker for evaluating end-product quality [83,84]. A conspicuous phenotype for mutants defective in SSP folding, assembly, and ER exit is the formation of abnormal ER-derived organelles called the MAG body in Arabidopsis and the ER-derived electron-dense PB in rice [75,85].…”