Proteomic exploration reveals a metabolic rerouting due to low oxygen during controlled germination of malting barley (Hordeum vulgare L.)

Abstract

Barley (Hordeum vulgare L.) is used in malt production for brewing applications. Barley malting involves a process of controlled germination that modifies the grain by activating enzymes to solubilize starch and proteins for brewing. Initially, the grain is submerged in water to raise grain moisture, requiring large volumes of water. Achieving grain modification at reduced moisture levels can contribute to the sustainability of malting practices. This study combined proteomics, bioinformatics, and biochemical phenotypic analysis of two malting barley genotypes with observed differences in water uptake and modification efficiency. We sought to reveal the molecular mechanisms at play during controlled germination and explore the roles of protein groups at 24 h intervals across the first 72 h. Overall, 3,485 protein groups were identified with 793 significant differentially abundant (DAP) within and between genotypes, involved in various biological processes, including protein synthesis, carbohydrate metabolism, and hydrolysis. Functional integration into metabolic pathways, such as glycolysis, pyruvate, starch and sucrose metabolism, revealed a metabolic rerouting due to low oxygen enforced by submergence during controlled germination. This SWATH-MS study provides a comprehensive proteome reference, delivering new insights into the molecular mechanisms underlying the impacts of low oxygen during controlled germination. It is concluded that continued efficient modification of malting barley subjected to submergence is largely due to the capacity to reroute energy to maintain vital processes, particularly protein synthesis.

Advancing Sustainable Malting Practices: Aquaporins as Potential Breeding Targets for Improved Water Uptake during Controlled Germination of Barley (Hordeum vulgare L.)

“…Most of the processes involved in germination are inseparable from AQP-facilitated water uptake in seeds, ,, also observed in our previous study . Due to the importance of barley in agriculture, several studies have investigated the role of AQPs in germination in field environments. ,,, However, information about their role in imbibition during controlled germination for malt barley production is limited.…”

“…In this study, we explored the proteome of two malting barley genotypes with demonstrated differences in water uptake during the initial stages of controlled germination. In previous research, IGB1467 demonstrated efficient proteolysis requiring less moisture (correlating with less water uptake), a desirable malting phenotype for improving the sustainability of malting practices . Our results found that upon imbibition, both IGB1467 and Flinders showed significant changes in tissue water content (Figures and S1).…”

“…Seven AQP proteins with at least two unique peptides per protein were selected for MRM validation analysis to confirm their abundance patterns and potential contribution to water uptake (Figure and Supporting Information Data S5B). Our SWATH-MS analysis showed an observed difference between the genotypes across the time points (Figure ). Similar patterns were observed in IGB and FLN for PIP1;1, PIP2;4, TIP2;3 TIP3;1, and TIP3;2 (Figure A,C,E,F,G, respectively), with an initial increase from 0 to 24 HAI followed by a decrease in abundance from 24 to 48 HAI.…”

“…To illustrate the functional differences of AQP abundance between IGB and FLN that may contribute to the increased grain modification efficiency seen in IGB, we explored the fold change (log 2 ) values between time points (0/24 and 24/48 HAI) of TIPs affecting vacuolation for storage reserve mobilization (Supporting Information Data S7). We included the changes also observed in the precursor proteins for abscisic acid (ABA) and gibberellic acid (GA) biosynthesis, given their modulation of TIPs , (Figure , Supporting Information Datas S1, S2, and S8).…”

“…We saw earlier changes in protein abundance, and a larger number of DAPs were identified with an increased abundance (Figure ). This observation is relevant to malting practices, contributing to the previously described malting phenotypes, and supports the hypothesis that IGB1467 exhibits significantly less water uptake in the first 24 HAI while demonstrating efficient modification for greater germination vigor at lower moisture content than Flinders.…”