Production of active angiotensin-I converting enzyme inhibitory peptides derived from bovine beta-casein by recombinant DNA technologies.
A wide range of anti-hypertensive peptides potentially able to lower blood pressure through the inhibition of vasoactive enzymes such as angiotensin-I converting enzyme (ACE) are known. Currently, ACE-inhibitory peptides can be produced from precursor proteins via enzymatic hydrolysis by proteolytic enzymes, or food fermentation with proteolytic starter cultures. These approaches are neither selective nor easy. In this study a novel procedure has been developed, based on recombinant DNA technologies, for the production of highly purified fractions of three polypeptides derived from bovine beta-casein active as ACE inhibitors in vitro. The procedure includes peptide expression in Escherichia coli cells as recombinant fusion proteins, purification by affinity chromatography, cleavage by proteinase from a selected strain of Lactobacillus helveticus and isolation of bioactive peptides (BPs). The reported concentration of inhibitor needed to reduce at 50% ACE activity (IC(50)) values for single BP calculated in inhibiting the ACE enzyme gave results in agreement with the same parameters available in literature for other milk-derived BPs. This procedure could be used to obtain quantities of pure peptides to determine their interactions with ACE, with the aim of designing peptides that have stronger inhibitory properties and exhibit new pharmacological profiles. Moreover, its scale up would be of commercial interest for the production of functional foods, e.g., milk beverages with blood pressure-lowering effects.
Losacco
Maurizio
M
Department of Biochemistry and Molecular Biology, University of Bari, Bari, Italy.
Gallerani
Raffaele
R
Gobbetti
Marco
M
Minervini
Fabio
F
De Leo
Francesca
F
D000806:Angiotensin-Converting Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002364:Caseins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007778:Lactobacillus / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
D000806:Angiotensin-Converting Enzyme Inhibitors; D002364:Caseins; D004274:DNA, Recombinant; D010455:Peptides; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase