Article

Abstract

-The proteolytic system of lactic acid bacteria has been characterised in detail and numerous modified strains with null or increased specific proteolytic activities have been constructed or identified among natural strains. Based on this knowledge, our objective was to ferment milk with modified strains and produce mixtures of peptides with specific features corresponding to potential bio-activities. We used a collection of Lactococcus lactis negative mutants for peptidase activities available in the laboratory to ferment the milk. In particular, we focused our work on mutants lacking either aminopeptidase N, X-prolyl dipeptidyl aminopeptidase or tripeptidase in order to test their ability to form peptides with immunomodulating or antihypertensive activities. At the end of fermentation, supernatants were fractionated by RP-HPLC. Each fraction collected was analysed by Maldi-Tof MS and sequencing. We observed that mutants accumulated specific peptides consistent with the specificity of the missing peptidases. Some of the peptides identified present similarities with peptides having immunomodulating or anti-hypertensive effects. One of these was quantified. At the same time, we observed that the inhibition of angiotensin converting enzyme was stronger in supernatants obtained with mutant strains than in supernatant obtained using the wild-type strain. In conclusion, we showed that in some cases, modifications to the proteolytic system of Lactococcus lactis gave rise to significant differences in the mixtures of peptides produced during milk fermentation. The differences in bio-activity of these peptide mixtures were only partially determined in vitro and evidently need to be demonstrated in vivo. Exploitation of the biodiversity of the proteolytic system of lactic acid bacteria may enable a direct application of this work and undoubtedly a promising means of directly producing natural bio-active peptides in fermented milk products. Milk fermentation / Lactococcus lactis / bio-active peptide / proteolysis / mutantRésumé -Conséquences de modifications du système protéolytique de Lactococcus lactis sur l'accumulation de peptides potentiellement bio-actifs pendant la fermentation du lait. Le système protéolytique des bactéries lactiques a été caractérisé en détail et de nombreuses souches aux activités protéolytiques modifiées ont été construites ou identifiées parmi des souches sauvages. Notre objectif est de produire des mélanges de peptides ayant des activités biologiques, en utilisant ces souches modifiées pour fermenter le lait. Pour cela, nous avons utilisé une collection de mutants de Lactococcus lactis dont certaines activités peptidasiques ont été supprimées. En particulier, nous nous sommes intéressés à des souches n'ayant plus l'aminopeptidase N, la X-prolyl-dipeptidyl aminopeptidase, ou la tripeptidase pour tester leur aptitude à former des peptides ayant des activités * Corresponding author: algaron@jouy.inra.fr 116 F. Algaron et al.

κ-Casein as a source of short-chain bioactive peptides generated by Lactobacillus helveticus

“…The j-CN-derived sequence VIESPPEIN was produced by all the analyzed L. helveticus strains. This peptide was previously detected in milk fermented by a Lactococcus lactis wild-type strain (Algaron et al 2004). Similarly, Robert et al (2001) obtained the VIESPPEIN peptide using L. helveticus NCC 619 and confirmed the ACE inhibitory activity of this sequence.…”

Use of Mass Spectrometry to Profile Peptides in Whey Protein Isolate Medium Fermented by Lactobacillus helveticus LH-2 and Lactobacillus acidophilus La-5

“…Proline at the second position of a nonameric peptide also resulted in a dramatic drop of the OppA binding affinity (122). Phosphorylated peptides and peptides high in proline content are more resistant to hydrolysis by proteolytic enzymes (123, 124). Oligopeptides containing VPP and IPP sequences could release branched chain amino acids (BCAAs) in the presence of specific peptidases.…”

Effect of Organic Nitrogen Source on Angiotensin I Converting Enzyme (ACE) Inhibitory Peptides Fermented by <em>Lactobacillus bulgaricus</em> LB6 from Goat Milk

“…lactis, L. lactis ssp. Cremoris and Enterococcus faecalis (Algaron et al, 2004;Ashar and Chand, 2003;Fuglsang et al, 2003b;Gobbetti et al, 2000;Herna´ndez-Ledesma et al, 2004;Leclerc et al, 2002;Muguerza et al, 2006;Nakamura et al, 1995;Quiro´ s et al, 2007;Robert et al, 2004;Rodríguez-Figueroa et al, 2010;Rokka et al, 1997;Shuangquan and Taku, 2008;Vermeirssen et al, 2003;Yamamoto et al, 1994aYamamoto et al, , b, 1999. In our previous study, 28 probiotic Lactobacillus strains were used to ferment goat milk to screen for production ACE-inhibitory peptides, the results showed that 20 strains had ACE inhibitory activity and among them of 4 strains including Lactobacillus reuteri, Lactobacillus bulgaricus, Lactobacillus rhamnosus and Lactobacillus helveticus were especially significant as producers of ACE-inhibitory peptides (He et al, 2012).…”