Screening for nutritive peptides that modify cholesterol 7alpha-hydroxylase expression.
Bioactive peptides with a variety of effects have been described from several nutritive proteins. They exhibit antimicrobial, blood-pressure lowering, antithrombotic, immunomodulatory, and cholesterol-modulating effects. In this study, we have examined whether peptides derived from food proteins might influence bile acid synthesis. A reporter gene cell line that carries a cholesterol 7alpha-hydroxylase promoter fragment fused to firefly luciferase ( cyp7a-luc) was used to screen for nutritive peptides affecting cyp7a expression, the enzyme catalyzing the rate-limiting step in bile acid synthesis. Proteolytic hydrolysates were prepared from soy protein and bovine casein with pepsin, trypsin, chymotrypsin, and elastase and size fractionated using ultrafiltration. Several bioactive hydrolysates could be identified that inhibited luciferase expression. Also, an activation of kinase (AKT, ERK, p38-MAPK) signaling could be observed. Selected hydrolysates were further fractionated by reversed-phase HPLC. Bioactive HPLC-fractions were obtained from casein but not from soy hydrolysates; however, activity could not be recovered in single peak fractions. Peptides in such fractions were identified by mass spectrometry. Five selected peptides from alpha S1-casein present in active fractions were synthesized, but none of these showed activity in the cyp7a-luc screening system. However, two of them activated MAP-kinase signaling similar to the hydrolysates, which suggests, that these peptides are involved in cyp7a regulation by the casein hydrolysates.
Nass
Norbert
N
Department of Cardiothoracic Surgery, Martin Luther University Halle-Wittenberg, Ernst-Grube Str.40, D-06120 Halle/Saale, Germany. norbert.nass@medizin.uni-halle.de
Schoeps
Regina
R
Ulbrich-Hofmann
Renate
R
Simm
Andreas
A
Hohndorf
Lars
L
Schmelzer
Christian
C
Raith
Klaus
K
Neubert
Reinhard H H
RH
Eder
Klaus
K
D000595:Amino Acid Sequence; D000818:Animals; D001647:Bile Acids and Salts / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002364:Caseins / Q000737:chemistry / Q000494:pharmacology; D002417:Cattle; D045744:Cell Line, Tumor; D002784:Cholesterol / Q000378:metabolism; D002790:Cholesterol 7-alpha-Hydroxylase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects*; D017930:Genes, Reporter; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000494:pharmacology*; D011492:Protein Hydrolysates / Q000737:chemistry / Q000494:pharmacology; D030262:Soybean Proteins / Q000737:chemistry / Q000494:pharmacology
D001647:Bile Acids and Salts; D002364:Caseins; D010455:Peptides; D011492:Protein Hydrolysates; D030262:Soybean Proteins; C475729:casein hydrolysate; D002784:Cholesterol; D002790:Cholesterol 7-alpha-Hydroxylase; D020928:Mitogen-Activated Protein Kinases