Transportation of whey protein-derived peptides using Caco-2 cell model and identification of novel cholesterol-lowering peptides.

Background: The increasing morbidity and mortality of cardiovascular disease have become a major factor in human death. Serum cholesterol is considered to be an important risk factor for inducing coronary heart disease, atherosclerosis and other cardiovascular diseases. To screen intestinal absorbable functional small peptides with cholesterol-lowering activity by enzymatic hydrolysis of whey protein and develop cholesterol-based functional food that may become a substitute for chemically synthesized drugs, providing new ideas for diseases caused by high cholesterol. Objective: This study aimed to evaluate the cholesterol-lowering activity of intestinal absorbable whey protein-derived peptides hydrolyzed by alkaline protease, trypsin and chymotrypsin, respectively. Method: The whey protein hydrolysates acquired by enzymatic hydrolysis under optimal conditions were purified by a hollow fiber ultrafiltration membrane with a molecular weight cutoff of 10 kDa. The fractions obtained by Sephadex G-10 gel filtration chromatography were transported through a Caco-2 cell monolayer. The transported peptides were detected in the basolateral aspect of Caco-2 cell monolayers using ultra- performance liquid chromatography-tandem mass spectrometry (UPLC-MS). Results: His-Thr-Ser-Gly-Tyr (HTSGY), Ala-Val-Phe-Lys (AVFK) and Ala-Leu-Pro-Met (ALPM) were unreported peptides with cholesterol-lowering activity. The cholesterol-lowering activities of the three peptides did not change significantly during simulated gastrointestinal digestion. Conclusion: This study not only provides theoretical support for the development of bioactive peptides that can be directly absorbed by the human body, but also provides new treatment ideas for hypercholesterolemia.

Identification and Characterization of Domains Responsible for Cell Wall Binding, Self-Assembly, and Adhesion of S-layer Protein from Lactobacillus acidophilus CICC 6074.

Lactobacillus S-layer protein (SLP) is a biologically active protein on the cell surface. To further elucidate the structures and functions of SLP in Lactobacillus acidophilus CICC 6074, this study was conducted to identify the functional domains of SLP which is responsible for cell wall anchoring, self-assembly, and adhesion. The gene (slpA) of L. acidophilus CICC 6074 SLP was amplified by polymerase chain reaction and speculated functional domains. Fusion proteins of C-terminal truncations from SLP were exogenously expressed in Escherichia coli BL21 (DE3). FITC-labeling N-terminal truncations of SLP were synthesized. The C-terminal domain was more likely to be the binding region, and the cell wall-anchored receptor of SLP was teichoic acid. Furthermore, N-terminal truncations could self-assemble to milk fat globule membrane polar lipid liposomes observed using a fluorescence microscope. Notably, SAN1 (region 32-55) of N-terminal truncations was mainly responsible for the adhesion of SLP to HT-29 cells. These results showed that SLP played a crucial role in the functions of L. acidophilus CICC 6074, which might be of significant reference value for future studies.