Purification, identification and molecular docking of dual-functional peptides derived from corn protein hydrolysates with antioxidant and antiadhesive activity against Helicobacter pylori.

BACKGROUND: More than 50% of the world's population is infected with Helicobacter pylori, which is classified as a group I carcinogen by the World Health Organization (WHO). RESULTS: Corn protein dual-functional peptides were identified and functionally analyzed in vitro and in silico. Two novel dual-functional peptides were identified as Cys-Gln-Asp-Val-Pro-Leu-Leu (CQDVPLL, CQ7) and Thr-Ile-Phe-Pro-Gln-Cys (TIFPQC, TI6) using nanoscale liquid chromatography coupled to tandem mass spectrometry (nano-LC-MS/MS). The antiadhesive effects against H. pylori of CQ7 and TI6 were 45.17 ± 2.41% and 48.62 ± 1.84% at 4 mg mL-1 , respectively. In silico prediction showed that CQ7 and TI6 had good physicochemical properties. Molecular docking demonstrated that CQ7 and TI6 could bind to the adhesins BabA and SabA by hydrophobic interactions and hydrogen bonds, preventing H. pylori infection. Moreover, CQ7 showed strong antioxidant activity due to its unique amino acid composition. CONCLUSION: The present study demonstrated that the identified peptides, CQ7 and TI6, possess antioxidant and antiadhesive effects, preventing H. pylori infection and alleviating oxidative injury to the gastric mucosa. © 2023 Society of Chemical Industry.

Preparation of Corn Peptides with Anti-Adhesive Activity and Its Functionality to Alleviate Gastric Injury Induced by Helicobacter pylori Infection In Vivo.

More than 50% of the world population is infected with Helicobacter pylori (H. pylori), which is classified as group I carcinogen by the WHO. H. pylori surface adhesins specifically recognize gastric mucosal epithelial cells' (GES-1 cells) receptor to complete the adhesion. Blocking the adhesion with an anti-adhesion compound is an effective way to prevent H. pylori infection. The present study found that corn protein hydrolysate, hydrolyzed by Neutral, effectively alleviated gastric injury induced by H. pylori infection through anti-adhesive and anti-inflammatory effects in vitro and in vivo. The hydrolysate inhibited H. pylori adhesion to GES-1 cells significantly, and its anti-adhesive activity was 50.44 ± 0.27% at 4 mg/mL, which indicated that the hydrolysate possessed a similar structure to the GES-1 cells' receptor, and exhibited anti-adhesive activity in binding to H. pylori. In vivo, compared with the H. pylori infection model group, the medium and high dose of the hydrolysate (400-600 mg/kg·bw) significantly decreased (p < 0.05) the amount of H. pylori colonization, pro-inflammatory cytokines (IL-6, IL-1ß, TNF-α and MPO), chemokines (KC and MCP-1) as well as key metabolites of NF-κB signaling pathway levels (TLR4, MyD88 and NF-κB), and it increased antioxidant enzyme contents (SOD and GSH-Px) and the mitigation of H. pylori-induced pathological changes in the gastric mucosa. Taken together, these results indicated that the hydrolysate intervention can prevent H. pylori-induced gastric injury by anti-adhesive activity and inhibiting the NF-κB signaling pathway's induction of inflammation. Hence, the corn protein hydrolysate might act as a potential anti-adhesive agent to prevent H. pylori infection.

Conjugation of the glutelin hydrolysates-glucosamine by transglutaminase and functional properties and antioxidant activity of the products.

A novel mixture of glycopeptides was prepared from corn glutelin and glucosamine (GlcN). The functional properties and antioxidative activities of this mixture were investigated. Corn glutelin was limited hydrolyzed by Alcalase, and then its hydrolysates were glycosylated with GlcN by transglutaminase (TGase) to modify its main and side chain, respectively. Under the optimized conditions, the content of GlcN conjugated to peptides was 81.98 ± 1.98 mg/g glutelin peptides. According to electrospray ionization mass spectrometry (ESI-MS) analysis, there are two types of glycopeptides in the mixture, TGase and non-enzymatic glycated counterparts. Compared with original glutelin, the glycosylated glutelin hydrolysates exhibited better solubility in the pH range of 2-11 and other functional properties except foaming stability. Meanwhile, it is more easily digested by pepsin and trypsin, and possessed excellent antioxidative activities. It also exhibited cytoprotective effects and intracellular ROS scavenging activities in LO2 cells subjected to oxidative stress by oxidation with ethanol solution.

Gene cloning, expression and homology modeling of first fibrinolytic enzyme from mushroom (Cordyceps militaris).

Fibrinolytic enzymes are important thrombolytic agents for blood-clotting disorders like cardiovascular diseases. Availability of novel recombinant fibrinolytic enzymes can overcome the shortcomings of current thrombolytic drugs. With the objective of facilitating their cost-effective production for therapeutic applications and for gaining deeper insight into their structure-function, we have cloned and expressed the first fibrinolytic protease gene from Cordyceps militaris. Cordyceps militaris fibrinolytic enzyme (CmFE) has one open reading frame of 759 bp encoding "pre-pro-protein" of 252 amino acids. Recombinant CmFE was expressed as 28 kDa extracellular enzyme in Pichia pastoris which was capable of degrading fibrin clot. A structure homology model of CmFE was developed using urokinase-type plasminogen activator. The active site contains catalytic triad His41, Asp83, Ser177 and consensus sequence of GDSGG. The substrate binding residues are Asp (171), Gly (194) and Ser (192). Its trypsin-like specificity is determined by the critical Asp171 in S1 subsite. The "oxyanion hole" is formed by backbone amide hydrogen atoms of Gly-175 and Ser-177. CmFE contains six conserved cysteines forming three disulfide linkages. This is the first study describing cloning, expression and prediction of structure-function relationship of a mushroom fibrinolytic protease. Hence it has great relevance in application of fibrinolytic enzymes as thrombolytic agents.

Preparation of antioxidative corn protein hydrolysates, purification and evaluation of three novel corn antioxidant peptides.

Corn gluten meal is a major co-product of corn wet milling. Corn gluten meal was hydrolyzed with Alcalase, Flavourzyme, Alcalase+Flavourzyme and Flavourzyme+Alcalase. At the substrate concentration of 10%, corn protein hydrolysate catalyzed by Alcalase had a degree of hydrolysis of 17.83%, which was higher than that by Flavourzyme (3.65%). The hydrolysate catalyzed by Alcalase+Flavourzyme exhibited better antioxidant activities and was further purified. Three novel antioxidant peptides were purified by a series of chromatographic techniques. Sequences of the three peptides were identified as Cys-Ser-Gln-Ala-Pro-Leu-Ala, Tyr-Pro-Lys-Leu-Ala-Pro-Asn-Glu and Tyr-Pro-Gln-Leu-Leu-Pro-Asn-Glu, respectively. Among the three peptides, Cys-Ser-Gln-Ala-Pro-Leu-Ala exhibited good reducing power and excellent scavenging capacities for DPPH radical and superoxide anion radical, with IC50 values of 0.116 and 0.39mg/ml, respectively. The results from our study indicate antioxidant potency of corn protein hydrolysates and peptides separated from corn gluten meal and can provide basic understanding for the application of corn protein hydrolysates as natural antioxidants.

Production of hydrolysate with antioxidative activity by enzymatic hydrolysis of extruded corn gluten.

Hydrolysate of extruded corn gluten with higher solubility and antioxidative property was prepared. Extrusion and starch removal of corn gluten were applied as pretreatment before enzymatic hydrolysis by Alcalase. The amylase hydrolysis of starch at 70 degrees C for 3 h resulted in the removal of the starch from the extruded corn gluten. The best hydrolysis results can be obtained by conducting the hydrolysis at 60 degrees C with water addition 20 g/g protein, enzyme addition 0.048 Ansen units/g protein, pH 8.5, and 120 min. Degree of hydrolysis of extruded and nonextruded corn gluten reached 39.54 and 31.16%, respectively, under the optimal condition. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the optimal hydrolysate revealed that proteolysis of extruded corn gluten was more extensive than proteolysis of its counterpart which was not subjected to extrusion. The molecular weight of the peptides in the optimal hydrolysate was mainly over 3,710-660 Da as determined by gel filtration chromatography. The hydrolysates displayed good solubility and antioxidative activity. The separation profile of the hydrolysate on an ion exchange chromatography of Q-Sepharose Fast Flow showed that many kinds of peptides had antioxidative effect. A new peptide with antioxidative activity was purified, and its amino acid sequence was Phe-Pro-Leu-Glu-Met-Met-Pro-Phe, which was identified by Q-TOF2 mass spectrometry.