Screening Of Xanthine Oxidase Inhibitor Peptides From Scomberomorus Niphonius Protein Based On Ligand Fishing

Xanthine oxidase?XOD?is the last enzyme of purine metabolism in the human body,treating hyperuricemia by inhibiting xanthine oxidase activity is one of the most important methods to prevent gout.The purpose of this article is to obtain bioactive peptides having a xanthine oxidase inhibitory effect from the complex Scomberomorus niphonius protein hydrolyzate,based on"ligand fishing".Purine content was measured by HPLC to verify the peptide's inhibitory effect on xanthine oxidase.This experiment tested the type of inhibition and antioxidant capacity to explain the mechanism of inhibition.The results of this paper were as follows:1.Four kinds of purines in aquatic products were detected by high performance liquid chromatography.The linear relationship,precision,repeatability and recovery ability results showed that four purines could be completely separated before 11 min.Purine content in various parts of the common marine shellfish tissue was measured.And the highest purine content in shellfish was Patinopecten yessoensis,the richest tissue was the gland,up to 148.09 mg/100g.Neptunea cumingii had the highest purine content in snails,and also the highest part was the gland,up to 155.88 mg/100g.So we recommend that people with hyperuricemia should avoid eating these.2.Magnetic nanoparticles were prepared by the sol-gel method.Use this as the kernel,and used 3-aminopropyltriethoxysilane to surface modification,the glutaraldehyde was used in the activation process.Xanthine oxidase was immobilized on the surface of the silica magnetic particles.Infrared spectroscopy,XRD and TEM were used to characterize magnetic nanoparticles.It was observed that the silica could be uniformly distributed on the surface of magnetic beads,and xanthine oxidase was immobilized on the surface of magnetic nanoparticles.The amount of fixed oxidase reached 116.12?g/mg.3.Then screened out xanthine oxidase inhibitor peptides from Scomberomorus niphonius protein hydrolysates based on ligand fishing by magnetic nanoparticles,got four amino acid sequences.The peptides were identified by HPLC-MS/MS.Two sequences,IIAPPER and AGFAGDDAPR,were synthesized for identification of inhibitory effects.IC₅₀ with which were 6.08 mg/mL and 6.15 mg/mL,respectively.4.The peptide IIAPPER was taken for the next study to evaluate its antioxidant capacity and the mechanism of its inhibition was preliminary discussed.Fluorescence spectrum results showed that the active peptide can quench xanthine oxidase fluorescence intensity.The DPPH scavenging ability was equivalent to 0.033 mM/mL of vitamin C and the total antioxidant capacity was equivalent to 0.76 mM/mL Trolox.ADME prediction results showed that the blood-brain barrier permeability of this active peptide was extremely low.According to the result of the docking of the binding molecule,it could enter the active site of xanthine oxidase inhibition,thereby inhibiting the conversion of xanthine and hypoxanthine to uric acid.Theoretically,the amino acid composition of IIAP has higher antioxidant capacity.However,the inhibition of xanthine oxidase by IIAP and PPER showed that IIAP was less effective than PPER.