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Lentil and fava bean with contrasting germination kinetics: a focus on digestion of proteins and bioactivity of resistant peptides

ABSTRACT: Germination offers advantages to improve legume protein digestibility as it disintegrates seed structure and hydrolyzes proteins and anti-nutrients. Seed permeability (related to polyphenol content of seed coats) is an important factor affecting the duration of seed germination and its impact in protein digestibility and bioactivity. The objective was to compare the effect of seed germination on protease activity, structure and proteolysis of four selected legumes with contrasting seed coat polyphenol profiles (gray zero tannin [GZL], beluga [BL] and dehulled red [DL] lentils; and zero tannin/low vicine-convicine fava bean [ZF]). Protein hydrolysis was characterized during germination and digestion with respect to proteins, peptides and free amino acids (FAA). In vitro antihypertensive and antioxidant activities of digests were investigated, and peptidomic characterization (HPLC-MS/MS) and identification of bioactive fragments in intestinal digests were performed. Regardless of seed type, germination increased protease activity and reduced levels of phytic acid, trypsin inhibitors and tannins (only in BL). Significant proteolysis of the 7S and 11S globulins and concomitant increase peptides and FAA was observed in all sprouted legumes. Digestion kinetics in sprouts revealed a faster generation of FAA and peptides than in dry seeds, with changes more evident for DL associated to faster imbibition, germination and sprout growth. In contrast, BL sprouts showed the lowest protein digestibility, likely due to lower protease activity levels, seed structure disintegration and higher anti-nutrient levels in comparison to GZL, DL and ZF. Moreover, digestion of sprouts resulted in a higher number of resistant peptides in DL and ZF that matched with previously reported bioactive sequences, suggesting a promising health potential of legume sprouts that was confirmed in vitro. Results suggested that the germination process improved protein digestibility and health promoting potential of lentil and fava bean proteins although these changes were more evident in DL due to its rapid imbibition, faster germination and sprout development. This study will provide important information for either plant breeders to develop legume varieties with permeable seed coats or food producers that could use dehulled seeds for efficient production of sprouts as sustainable food sources of plant proteins with improved nutritional and healthy properties.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Vicia Faba Lens Culinaris

TISSUE(S): Plant Cell

SUBMITTER: Cristina Martínez Villaluenga

LAB HEAD: Cristina Martínez-Villaluega

PROVIDER: PXD027847 | Pride | 2021-11-02

REPOSITORIES: Pride

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Project description:Purpose: The goals of this study are to analyze the transcriptome of five time point in broccoli seed germination and sprout development and to find the putative glucosinolate metabolism genes in the stage. Methods: Total mRNA of germinated seeds, 3 day cotyledons, 7 day botyledons, 11 day cotyledons and 11 day euphyllas of wild-type broccoli were harvested. Each sample was harvested in three independent biological replicates with equal weight and subsequently pooled together for sequencing. The sequence reads that passed quality filters were de novo assembled using VELVET followed by OASES. Then the assembled unigenes were used for the abundance and functional analysis. Results: A total of ~85million 251bp reads were obtained. After de novo assembly and searching the assembled transcripts against the Arabidopsis thaliana and Nr databases, 19,441 top-hit transcripts were clustered as unigenes with an average length of 2,133bp. These unigenes were classified according to their putative functional categories. Cluster analysis of total unigenes with similar expression patterns and differentially expressed unigenes among different tissues,as well as transcription factor analysis were performed. We identified 25 putative glucosinolate metabolismgenes sharing 62.04-89.72% nucleotide sequence identity with the Arabidopsis orthologs. This established a broccoli glucosinolate metabolic pathway with high colinearity to Arabidopsis. Many of the biosynthetic and degradation genes showed higher expression after germination than in seeds; especially the expression of the myrosinaseTGG2 was 20-130 times higher.These results along with the previous reports that glucosinolate concentration decreased exponentially once after germination indicate the breakdown products of glucosinolates may play important roles in broccoli seed germination and sprout development. Conclusion: Our study provides the largest genetic resource of broccoli to date. These data will pave the way for further studies and genetic engineering of broccoli sprouts to develop functional vegetables containing high levels of the anticarcinogenic glucosinolates. They will also provide new insight into the genomic research of this species and its relatives. Wild-type broccoli mRNA profiles of seeds, 3 day cotyledons, 7 day botyledons, 11 day cotyledons and 11 day euphyllas were generated by deep sequencing, three biological replicates pooling together for each tissue, using Illumina Myseq platform.

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Lentil and fava bean with contrasting germination kinetics: a focus on digestion of proteins and bioactivity of resistant peptides

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