YebC, a putative transcriptional factor involved in the regulation of the proteolytic system of Lactobacillus

Lucía Brown,Fernanda Mozzi,Silvina Fadda,Mariano Elean,Elvira M Hebert,Lucila Saavedra,Josefina M Villegas

doi:10.1038/s41598-017-09124-1

Abstract

The proteolytic system of Lactobacillus plays an essential role in bacterial growth, contributes to the flavor development of fermented products, and can release bioactive health-beneficial peptides during milk fermentation. In this work, a genomic analysis of all genes involved in the proteolytic system of L. delbrueckii subsp. lactis CRL 581 was performed. Genes encoding the cell envelope-associated proteinase, two peptide transport systems, and sixteen peptidases were identified. The influence of the peptide supply on the transcription of 23 genes involved in the proteolytic system of L. delbrueckii subsp. lactis was examined after cell growth in a chemically defined medium (CDM) and CDM supplemented with Casitone. prtL, oppA1, optS, optA genes as well as oppDFBC and optBCDF operons were the most highly expressed genes in CDM; their expression being repressed 6- to 115-fold by the addition of peptides. The transcriptional analysis was confirmed by proteomics; the up-regulation of the PrtL, PepG, OppD and OptF proteins in the absence of peptides was observed while the DNA-binding protein YebC was up-regulated by peptides. Binding of YebC to the promoter region of prtL, oppA1, and optS, demonstrated by electrophoretic mobility shift assays, showed that YebC acts as a transcriptional repressor of key proteolytic genes.

Highlights

Lactobacillus (L.) delbrueckii subsp. lactis is a homofermentative thermophilic lactic acid bacterium (LAB) widely used as starter culture in dairy fermentation processes, such as fermented sour milks and Swiss- and Italian-type cheeses
L. delbrueckii subsp. lactis CRL 581, a strain isolated from a homemade Argentinian hard cheese, possesses a cell envelope-associated proteinase (CEP) (PrtL) that is able to release a series of bioactive health-beneficial peptides from α- and β-caseins11, 12
The present study was focused on the proteolytic system of L. delbrueckii subsp. lactis CRL 581, which plays an essential role in bacterial growth contributing to the organoleptic properties of fermented products; the release of bioactive health-beneficial peptides during milk fermentation can occur7, 12, 20

Summary

Introduction

Lactobacillus (L.) delbrueckii subsp. lactis is a homofermentative thermophilic lactic acid bacterium (LAB) widely used as starter culture in dairy fermentation processes, such as fermented sour milks and Swiss- and Italian-type cheeses. As the concentration of amino acids and small peptides in milk is very limited, the proteolytic system of LAB is crucial to obtain essential amino acids from caseins during growth in this substrate, thereby ensuring a successful fermentation This proteolytic system consists of (i) an extracellular cell envelope-associated proteinase (CEP) that hydrolyzes caseins into oligopeptides; (ii) specialized transport systems that take up peptides into the cell; and (iii) several intracellular peptidases, which degrade peptides into amino acids . This proteolytic system consists of (i) an extracellular cell envelope-associated proteinase (CEP) that hydrolyzes caseins into oligopeptides; (ii) specialized transport systems that take up peptides into the cell; and (iii) several intracellular peptidases, which degrade peptides into amino acids2, 8, 9 This proteolytic system contributes to the flavor and texture development of fermented products , and may release bioactive health-beneficial peptides during milk fermentation. Such an integrated approach can be used to predict the proteolytic potential of Lactobacillus to expand the knowledge on specific features likely related to its industrial applications

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