Marine-derived antioxidant peptide modified bodies with transmembrane activity and application of marine-derived antioxidant peptide modified bodies

Abstract

The invention relates to the field of biological medicines, in particular to a group of marine-derived antioxidant peptide modifiers with transmembrane activity and application thereof. According to the method, marine-derived antioxidant peptides are subjected to preliminary analysis and batch screening on the basis of a bioinformatics tool, amino acids of the antioxidant peptides are further modified according to the structure-function relationship of the antioxidant peptides, toxicity, transmembrane capacity and Keap1 protein binding capacity prediction are combined, and finally a group of high-activity antioxidant peptide modified bodies are obtained through activity verification. Compared with common glutathione, the antioxidant peptide modified body disclosed by the invention has stronger free radical scavenging capacity, can enter cells to influence an antioxidant signal channel and improve the antioxidant capacity of the cells, and has an obvious protective effect on cell oxidative stress injury induced by ultraviolet rays. The invention provides a screening modification process of a high-activity antioxidant peptide and a group of anti-ultraviolet radiation damage polypeptides with remarkable activity. The anti-ultraviolet radiation damage polypeptides have wide application prospects in the fields of antioxidant drugs, anti-ultraviolet damage protection and the like.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, in particular to a group of marine-derived antioxidant peptide modifications with transmembrane activity and applications thereof. Background Art

[0002] Long-term excessive exposure to ultraviolet radiation can induce an increase in reactive oxygen free radicals (ROS) in skin cells, especially ultraviolet rays (UVB), which can cause a variety of biological damages, leading to pigmentation, photoaging, and even skin cancer. The use of exogenous antioxidants can enhance the antioxidant capacity of the skin and effectively reduce skin damage caused by ultraviolet radiation. At present, the antioxidants commonly used in anti-ultraviolet radiation skin care products are mainly derived from artificial synthesis or natural plant extracts. Some of these antioxidants may have adverse pro-oxidative effects on the human body due to their different structures, dosages, and environments. Therefore, it is necessary to find more effective and safe natural antioxidants to improve the effectiveness and safety of anti-ultraviolet skin care products.

[0003] Antioxidant peptides are a class of bioactive peptides with antioxidant activity, usually composed of 2-20 amino acid residues. They have attracted attention because of their low toxicity and side effects, environmental friendliness, and ability to effectively scavenge reactive oxygen free radicals, target key proteins in the intracellular antioxidant signaling pathway, and significantly improve the antioxidant capacity of cells. Marine biological resources are rich, and they contain a large number of structurally specific and highly active substances. At present, a variety of peptides with significant antioxidant activity have been isolated from marine organisms, and both in vivo and in vitro studies have shown good antioxidant and UV damage prevention effects. Therefore, marine organisms have become an important source of new antioxidant and UV radiation active substances. Due to the different amino acid composition and arrangement order of antioxidant peptides, there are large differences in their antioxidant activity and mechanism of action. In order to screen out highly active candidate antioxidant peptides, the study of peptide structure-activity relationship is crucial. For the evaluation of antioxidant peptide activity, the commonly used chemical model method and cell biological method are relatively labor-intensive and resource-intensive, while bioinformatics tools can be used for batch screening and preliminary analysis and prediction of antioxidant peptides, which greatly promotes the discovery and application of highly active antioxidant peptides. Summary of the invention

[0004] The purpose of the present invention is to provide a group of marine-derived antioxidant peptide modifications with transmembrane activity and applications thereof.

[0005] In order to achieve the above object, the present invention adopts the following technical solutions:

[0006] The present invention uses marine antioxidant peptides as screening objects, and screens out a total of 366 antioxidant peptides by searching peptide and literature databases such as BIOPEP, PubMed, and Web of Science. The online prediction scoring tool AnOxPePred (https: / / services.healthtech.dtu.dk / services / AnOxPePred-1.0 / , 2023.3.22) was used to obtain the top 20 highly active antioxidant peptide sequences. In order to further improve its antioxidant activity, variable amino acids were predicted by alanine scanning, and antioxidant amino acids (histidine, methionine, cysteine, tryptophan, tyrosine) were used for replacement to screen out candidate modified peptides with higher activity prediction scores. The online tools ToxinPred (https: / / webs.iiitd.edu.in / raghava / toxinpred / index.html, 2023.4.22) and BChemRF-CPPred (http: / / comptools.linc.ufpa.br / BChemRF-CPPred / , 2023.4.22) were further used to predict toxicity and transmembrane ability, and molecular docking was used to predict its binding ability with Keap1 protein. Combined with in vitro activity detection, a group of highly active antioxidant peptide modifications with transmembrane ability were finally screened. Studies have shown that compared with commonly used glutathione, the modified body has stronger free radical scavenging activity and can efficiently penetrate the cell membrane and enter the cell. Compared with vitamin C, the modified peptide can significantly reduce the intracellular ROS level, more effectively increase the intracellular antioxidant enzyme GPX activity, and has a significant protective effect on ultraviolet-induced cell oxidative damage. It has good application potential in the development of antioxidant drugs and anti-ultraviolet radiation protection products.

[0007] Based on the above technical scheme, the first aspect of the present invention provides a group of marine-derived antioxidant peptide modifications with transmembrane activity, wherein the amino acid sequence of the antioxidant peptide modifications is shown in any one of SEQ ID NO:1-SEQ ID NO:3.

[0008] Antioxidant peptide modification WP5 (SEQ ID NO: 1):

[0009] Tryptophan-Proline-Tryptophan-Histidine-Tryptophan (Abbreviation: Trp-Pro-Trp-His-Trp, WPWHW)

[0010] Antioxidant peptide modification LW5 (SEQ ID NO: 2):

[0011] Leucine-tryptophan-histidine-tryptophan-histidine (English abbreviation: Leu-Trp-His-Trp-His, LWHWH)

[0012] Antioxidant peptide modification YY6 (SEQ ID NO: 3)

[0013] Tyr-Tyr-Pro-Tyr-Trp-Leu (abbreviation: Tyr-Tyr-Pro-Tyr-Trp-Leu, YYPYWL)

[0014] SEQ ID NO:4:

[0015] Tryptophan-Proline-Aspartic acid-Histidine-Tryptophan (Abbreviation: Trp-Pro-Asp-His-Trp, WPDHW)

[0016] SEQ ID NO:5:

[0017] Leucine-Tryptophan-Histidine-Threonine-Histidine (English abbreviation: Leu-Trp-His-Thr-His, LWHTH)

[0018] SEQ ID NO:6:

[0019] Tyrosine-Tyrosine-Proline-Tyrosine-Glutamine-Leucine (Abbreviation: Tyr-Tyr-Pro-Tyr-Gln-Leu, YYPYQL)

[0020] Furthermore, the antioxidant peptide modified body WP5, whose amino acid sequence is shown in SEQ ID NO: 1, is a linear small peptide with a molecular weight of 810.92 Da, composed of L-type amino acid residues, and is modified by replacing aspartic acid in the parent peptide of the amino acid sequence of SEQ ID NO: 4 with tryptophan;

[0021] Furthermore, the antioxidant peptide modified body LW5 having an amino acid sequence as shown in SEQ ID NO: 2 is a linear small peptide with a molecular weight of 777.89 Da, composed of L-type amino acid residues, and is modified by replacing the threonine in the parent peptide having an amino acid sequence as shown in SEQ ID NO: 5 with tryptophan;

[0022] Furthermore, the antioxidant peptide modified body YY6 with an amino acid sequence as shown in SEQ ID NO: 3 is a linear small peptide with a molecular weight of 904.04 Da, composed of L-type amino acid residues, and is modified by replacing glutamine in the parent peptide with an amino acid sequence as shown in SEQ ID NO: 6 with tryptophan.

[0023] Furthermore, the preparation method of the antioxidant peptide modification comprises the following steps: synthesizing the entire amino acid sequence of the polypeptide using a twelve-channel semi-automatic polypeptide synthesizer; desalting and purifying by high performance liquid chromatography (HPLC) reverse phase column chromatography, and determining that its purity is greater than 95%; and determining the amino acid sequence and molecular weight of the synthesized polypeptide by mass spectrometry (MS) analysis to obtain the antioxidant peptide modification.

[0024] The second aspect of the present invention provides a group of modified marine-derived antioxidant peptides with transmembrane activity as described above and their compositions for use in the preparation of products for scavenging free radicals.

[0025] Furthermore, the modified antioxidant peptide removes 2,2-azino-bis(3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt free radical (ABTS) and / or hydroxyl free radical (·OH).

[0026] The third aspect of the present invention provides a group of marine-derived antioxidant peptide modifications with transmembrane activity and their compositions for use in the preparation of products for resisting ultraviolet radiation and the oxidative damage caused by it.

[0027] Furthermore, the products for resisting ultraviolet radiation and the oxidative damage caused by it refer to products that protect human skin from damage by ultraviolet radiation in various ways.

[0028] Furthermore, the antioxidant peptide modification can effectively reduce UVB-induced oxidative stress damage in human epidermal keratinocytes (HaCaT cells), significantly reduce the levels of intracellular ROS and lipid peroxidation stress product malondialdehyde (MDA), and increase the activity of intracellular superoxide dismutase (SOD) and glutathione peroxidase (GPX).

[0029] Furthermore, the modified antioxidant peptide can enter cells, directly remove reactive oxygen free radicals and affect the cell antioxidant signaling pathway, thereby improving the cell antioxidant capacity and protecting the cells from ultraviolet radiation damage.

[0030] The advantages of the present invention are:

[0031] 1. After preliminary analysis and batch screening of marine antioxidant peptides based on database batch screening and bioinformatics tools, the present invention further modified its amino acids according to the structure-activity relationship of the antioxidant peptides, combined with the prediction of toxicity, transmembrane ability and Keap1 protein binding ability, and finally obtained a group of marine antioxidant peptides WP5, LW5 and YY6 through activity verification. Compared with the commonly used antioxidant peptide glutathione, the modified peptide of the present invention has a stronger free radical scavenging ability, can enter the cell to affect the antioxidant signaling pathway, improve the cell antioxidant capacity, and has a stronger protective effect on ultraviolet-induced cell oxidative stress damage. Compared with vitamin C, an antioxidant commonly used in cosmetics, the modified peptide more significantly increases the activity of the intracellular antioxidant enzyme GPX and effectively reduces the intracellular ROS level. The antioxidant peptide modified body has a small molecular weight, a simple structure, is easy to prepare and has a high efficiency transmembrane ability.

[0032] 2. The present invention provides a screening and modification process for highly active antioxidant peptides and a group of peptides with significant activity against ultraviolet radiation damage, which provide a new resource for the research and development of safer and more efficient natural antioxidant drugs and anti-ultraviolet radiation damage products, and have broad application prospects in the fields of antioxidant drugs and anti-ultraviolet damage protection. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 The screening and modification process of antioxidant peptide modifications.

[0034] Figure 2 AB in it is the HPLC and MS analysis report of the antioxidant peptide modified WP5.

[0035] Figure 3 AB in it is the HPLC and MS analysis report of the antioxidant peptide modified LW5.

[0036] Figure 4 AB in it is the HPLC and MS analysis report of the antioxidant peptide modified YY6.

[0037] Figure 5 AB in the expression is the antioxidant peptide modification and glutathione's scavenging activity against ABTS free radicals and ·OH free radicals.

[0038] Figure 6 This is a cytotoxicity test of antioxidant peptide modifications at different concentrations.

[0039] Figure 7 The transmembrane activity of the antioxidant peptide modification was detected using an ELISA instrument.

[0040] Figure 8 The transmembrane activity of the antioxidant peptide modifications was detected using confocal fluorescence microscopy.

[0041] Fig. 9 Figures AB show the morphology and survival rate of HaCaT cells after irradiation with different doses of UVB.

[0042] Fig.10 AF in it represents the effect of antioxidant peptide modification on the survival rate, MDA, ROS and antioxidant enzyme activity of HaCaT cells after UVB irradiation.

[0043] Fig.11 AC in the figure represents the effect of antioxidant modification on the expression of antioxidant signaling pathway proteins in cells. DETAILED DESCRIPTION

[0044] The specific implementation methods provided by the present invention are described in detail below in conjunction with the examples. The experimental methods used in the examples of the present invention are all conventional methods unless otherwise specified.

[0045] Example 1: Preparation of antioxidant peptide modifications

[0046] S1: Preparation of modified antioxidant peptides. According to the amino acid sequence described in claims 1-6, the entire sequence is synthesized using a 12-channel semi-automatic peptide synthesizer; desalting and purification is performed using HPLC reverse phase column chromatography; the sample that meets the purity requirement is concentrated and then packaged and freeze-dried;

[0047] S2: Identification of modified antioxidant peptides. Take a small amount of the finished peptides and use mass spectrometry to identify the amino acid sequence and molecular weight of the peptides.

[0048] The results are:

[0049] The sequence of the antioxidant peptide modified WP5 is shown in SEQ ID NO: 1, with a molecular weight of 810.92 Da, a linear small peptide, and a purity greater than 95% ( Figure 2 The sequence of the modified antioxidant peptide LW5 is shown in SEQ ID NO: 2, with a molecular weight of 777.89 Da, a small linear peptide, and a purity greater than 95% ( Figure 3 The sequence of the modified antioxidant peptide YY6 is shown in SEQ ID NO: 3, with a molecular weight of 904.04 Da, a small linear peptide, and a purity greater than 95% ( Figure 4 ).

[0050] Example 2: In vitro free radical scavenging activity test of antioxidant peptide modifications

[0051] 1. ABTS free radical scavenging activity.

[0052] The modified antioxidant peptide and the commonly used antioxidant peptide drug glutathione in clinical practice were dissolved in deionized water to a concentration of 0.25 mg / mL. The total antioxidant capacity (T-AOC) detection kit (ABTS method) (purchased from Shanghai Biotechnology) was used for determination, and the detection was performed according to the operating steps in the instruction manual.

[0053] The experimental results are as follows Figure 5 As shown in A, at the sample concentration tested, the antioxidant peptide modifications WP5, LW5 and YY6 had scavenging activities against ABTS free radicals of 71.12±3.58%, 67.63±1.65%, 68.51±0.54%, respectively, and the scavenging activity of glutathione against ABTS free radicals was 61.51±1.02% (p<0.05). Compared with glutathione, the antioxidant peptide modifications had stronger scavenging activity against ABTS free radicals.

[0054] 2. OH ​​radical scavenging activity.

[0055] The modified antioxidant peptide and glutathione were dissolved in deionized water to a concentration of 1.5 mg / mL. The OH free radical scavenging ability detection kit (micro method) (purchased from Shanghai Biotechnology) was used for determination, and the detection was carried out according to the operating steps in the instruction manual.

[0056] The experimental results are as follows Figure 5 As shown in B, at the tested sample concentrations, the scavenging activities of the antioxidant peptide modifications WP5, LW5 and YY6 against hydroxyl radicals were 52.15±1.99%, 51.25±1.29%, 53.06±2.23%, respectively, and the scavenging activity of glutathione against hydroxyl radicals was 25.62±2.51% (p<0.001). The antioxidant peptide modifications were significantly better than glutathione in scavenging ·OH free radicals.

[0057] Example 3: Detection of cytotoxicity and transmembrane ability of modified antioxidant peptides

[0058] 1. Experimental Methods

[0059] (1) Cytotoxicity assay. HaCaT cells were inoculated in a high-glucose DMEM medium containing 10% fetal bovine serum and 1% double-antibody (penicillin and streptomycin). When the cells grew to more than 80% of the surface adhesion, the cells were collected and 100 μL of the cell suspension was inoculated into a 96-well plate (approximately 2×10 4The cells were placed in a well (cells / well), and the medium was aspirated after complete attachment. Serum-free high-glucose DMEM medium was added to the control group, and serum-free high-glucose DMEM medium containing different concentrations of antioxidant peptide modifications (0.5 mg / mL, 0.25 mg / mL, 0.125 mg / mL, 0.063 mg / mL, 0.031 mg / mL) was added to the sample group. After culturing for 24 hours, 10 μL of CCK-8 reagent was added, and a blank control group (serum-free high-glucose DMEM medium and 10 μL of CCK-8 reagent were added to the wells without cells). After incubation for 2 hours, the OD was detected using a multifunctional microplate reader (Varioskan LUX). 450 ; Cell survival rate (%) = (sample group OD值 -Blank Group OD值 ) / (control group OD值 -Blank Group OD值 ).

[0060] (2) Transmembrane ability test. Cells were plated in 24-well plates (about 1×10 5 cells) or 96-well plates (about 2×10 cells per well) 4 The cells were cultured in 24-well plates (10 cells / well), and after complete attachment, they were rinsed twice with phosphate buffered saline (DPBS). 500 μL (24-well plate) and 100 μL (96-well plate) of serum-free high-glucose DMEM medium containing fluorescein isothiocyanate (FITC)-labeled antioxidant peptide modifications (0.125 mg / mL) were added to the 24-well plate and 96-well plate, respectively, and the culture was continued for 24 hours; the cells were washed three times with DPBS; the fluorescence intensity of the 96-well plate was detected by a multifunctional microplate reader; the 24-well plate was fixed with 4% paraformaldehyde for 10 minutes, then stained with Hoechst33342, and detected by confocal microscopy.

[0061] 2. Experimental results

[0062] like Figure 6 As shown, after the cells were treated with the antioxidant peptide modification at a concentration below 0.5 mg / mL, the cell viability was above 80%, indicating that the antioxidant peptide modification had no obvious toxicity to the cells within this concentration range;

[0063] like Figure 7 As shown in the figure, compared with the control group, the fluorescence intensity of cells incubated with FITC-labeled (green labeled) antioxidant peptide modifications was significantly increased, indicating that the antioxidant peptide modifications can pass through the cell membrane and be localized in the cells; confocal microscopy also showed green fluorescence around the cell nucleus (stained blue part), indicating that the antioxidant peptide modifications are localized in the cells ( Figure 8 ).

[0064] Example 4: Protective effect of modified antioxidant peptides on ultraviolet radiation-induced cellular oxidative stress

[0065] 1. Experimental Methods

[0066] (1) Establishment of UVB-induced cellular oxidative stress model. HaCaT cells were seeded in 96-well plates (approximately 2×10 4 After the cells were completely attached to the wall, the medium was aspirated and washed once with DPBS, leaving a little DPBS at the bottom of the well plate. HaCaT cells were irradiated with UVB lamp (model: TL 20W / 01RS SLV / 25, PHILIPS) at a dose of 5 mJ / cm 2 、10mJ / cm 2 、15mJ / cm 2 、20mJ / cm 2 、25mJ / cm 2 The optimal irradiation dose was determined when the cell survival rate was maintained at 50%-70% after irradiation and incubation for 24 hours, and subsequent experiments were carried out at this dose.

[0067] (2) Protective effect of modified antioxidant peptides on UV radiation-induced cellular oxidative stress. As described above, HaCaT cells were inoculated in 96-well plates (about 2×10 4 cells), using UVB (the optimal irradiation dose determined above is 10 mJ / cm 2 ) to irradiate the cells. Immediately after irradiation, peptide incubation was performed. The blank control group (not irradiated, Control) and the model group (UVB) were added with 100 μL of serum-free high-glucose DMEM medium, the antioxidant peptide modified body group was added with serum-free high-glucose DMEM medium containing 0.25 mg / mL WP5, LW5 and YY6, and the positive control group was added with the same concentration of vitamin C (Vc) and glutathione. After culturing for 24 hours, 10 μL of CCK-8 reagent was added to each group, incubated for 2 hours, and the OD value at a wavelength of 450 nm was detected using an enzyme reader, and the cell survival rate was calculated.

[0068] (3) Determination of intracellular ROS content. Cells were seeded in 24-well plates (about 1×10 5 cells), using UVB (irradiation dose 10mJ / cm 2) to irradiate the cells. Immediately after irradiation, peptides were added for incubation. The blank control group and model group were added with 500 μL of serum-free high-glucose DMEM medium, the antioxidant peptide modified group was added with serum-free high-glucose DMEM medium containing 0.25 mg / mL WP5, LW5 and YY6, and the positive control group was added with the same concentration of Vc. The culture was continued for 24 hours; the medium was aspirated, and DPBS containing 10 μM ROS fluorescent probe 2,7-dichlorofluorescein diacetate (DCFH-DA) was added, and incubated at 37°C in the dark for 30 minutes; then the fluorescence microscope was used for detection, and the fluorescence intensity was analyzed using Image J software.

[0069] (4) Determination of intracellular MDA content. Cells were seeded in 6-well plates (about 3×10 5 After the cells were completely attached to the wall, the medium was aspirated and the cells were washed once with DPBS, leaving a little DPBS at the bottom of the well plate. 2 ) The cells were irradiated. Immediately after irradiation, the peptides were added for incubation. The blank control group and the model group were added with 2 mL of serum-free high-glucose DMEM medium, and the antioxidant peptide modified group was added with 2 mL of serum-free high-glucose DMEM medium containing 0.25 mg / mL WP5, LW5 and YY6. The positive control group was added with the same concentration of Vc and cultured for 24 hours. The cells were collected and tested according to the instructions of the MDA detection kit.

[0070] (5) Determination of intracellular antioxidant enzyme SOD and GPX activity. Cell inoculation, UVB irradiation and peptide treatment were the same as described in method (4). After the cells were collected, the SOD and GPX detection kits were followed for detection.

[0071] 2. Experimental results

[0072] like Fig. 9 As shown in A and 9B, the cell survival rate of HaCaT cells was significantly decreased after UVB irradiation (p<0.001).

[0073] Immediately after irradiation, the antioxidant peptide modification was added and incubated for 24 hours. Compared with glutathione, it could significantly increase the cell survival rate (p<0.01) and had a similar effect to that of Vc ( Fig.10 A).

[0074] After UVB irradiation, the levels of intracellular MDA and ROS increased significantly, indicating that the cells were in a state of oxidative stress; after treatment with antioxidant peptides and Vc, intracellular MDA ( Fig.10 B) and ROS levels, and the antioxidant peptide modified YY6 has a stronger effect on reducing ROS levels than Vc ( Fig.10C, 10D).

[0075] After UVB irradiation, the activities of intracellular antioxidant proteases SOD and GPX decreased significantly. Antioxidant peptide modification and Vc treatment could significantly increase the activity of intracellular SOD ( Fig.10 E) and GPX activity (p<0.001), and the effects of the three antioxidant modifications on improving GPX activity were significantly stronger than those of Vc ( Fig.10 F)(p<0.01).

[0076] Example 5: Mechanism of action of modified antioxidant peptides in improving cellular antioxidant capacity

[0077] 1. Experimental Methods

[0078] (1) Cell protein extraction. Cell inoculation, UVB irradiation and polypeptide treatment were the same as those described in Example 4, method (4). After 24 h of cell culture, the culture medium was aspirated and the cells were washed three times with 4°C precooled PBS buffer. The PBS was aspirated and 80 μL of precooled protein lysis buffer (containing 1% protease inhibitor PMSF and 1% phosphatase inhibitor) was added to each well. After lysis on ice for 5 min, the cells were collected into a 1.5 mL centrifuge tube and lysed for 15 min at 4°C. After sufficient lysis, the cells were centrifuged at 12,000×g for 20 min and the supernatant was taken for later use. The protein concentration was detected using a BCA protein assay kit (purchased from Thermo Fisher).

[0079] (2) Intracellular protein expression detection. Add loading buffer to the protein sample at a ratio of 4:1, and boil in a water bath for 5 minutes to fully denature the protein; use 10% SDS-PAGE for protein electrophoresis; after the electrophoresis, remove the gel plate, carefully cut the gel containing the target band with a gel cutting plate, place it in pre-cooled transfer buffer, and transfer the protein to the PVDF membrane; after the transfer, soak the PVDF membrane in 5% skimmed milk powder for 2 hours; place the PVDF membrane in the diluted primary antibody solution, incubate at room temperature for 2 hours, wash the membrane three times with washing buffer TBST, add the diluted secondary antibody solution, incubate at room temperature for 1 hour, wash the membrane again with TBST three times, take pictures with a gel chemiluminescence imager, and use Image J software for grayscale value analysis.

[0080] 2. Experimental results

[0081] like Fig.11 As shown in the figure, under normal circumstances, the expression levels of nuclear transcription factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in the intracellular antioxidant signaling pathway are relatively low. After UVB irradiation, the expression levels of Nrf2 and HO-1 increase to counteract the oxidative stress state of cells.

[0082] Compared with the UVB irradiation group, the antioxidant peptide modified WP5 and LW5 could significantly increase the expression levels of Nrf2 and HO-1 (p<0.05), suggesting that the two may further enhance the ability of cells to resist oxidative stress by acting on the antioxidant signaling pathway. The antioxidant peptide modified YY6 showed no statistical difference compared with the model group (p>0.05), suggesting that it may play an antioxidant role in cells through other mechanisms such as direct removal of ROS.

[0083] In summary, the group of marine-derived antioxidant peptide modifications described in the present invention have strong antioxidant activity. Compared with commonly used glutathione, the modified peptide has a stronger free radical scavenging ability and has a stronger protective effect on ultraviolet-induced cell oxidative damage. Compared with vitamin C, the modified peptide more significantly increases the activity of the intracellular antioxidant enzyme GPX, reduces the level of intracellular ROS, and effectively improves the cell oxidative stress damage caused by ultraviolet radiation. Its potential mechanism may be to directly scavenge free radicals by entering the cell, act on the antioxidant pathway, etc. The antioxidant peptide modification of the present invention has a small molecular weight, a simple structure and is easy to prepare. It has great potential for preventing and treating ultraviolet damage to the skin and has practical application and promotion value.

[0084] The preferred embodiments of the present invention have been specifically described above, but the present invention is not limited to the embodiments. Those skilled in the art may make various equivalent modifications or substitutions without violating the spirit of the present invention. These equivalent modifications or substitutions are all included in the scope defined by the claims of this application.

Claims

1. A group of marine-derived antioxidant peptide modifications with transmembrane activity, characterized in that: The amino acid sequence of the antioxidant peptide modification is shown in any one of SEQ ID NO: 1-SEQ ID NO:

3.

2. The antioxidant peptide modification according to claim 1, characterized in that: The antioxidant peptide modification is composed of L-type amino acid residues.

3. The antioxidant peptide modification according to claim 1, characterized in that: The antioxidant peptide modification is a linear chain.

4. The antioxidant peptide modification according to claim 1, characterized in that: When the amino acid sequence is as shown in SEQ ID NO: 1, the molecular weight of the antioxidant peptide modification is 810.92 Da and the isoelectric point is 7.80; when the amino acid sequence is as shown in SEQ ID NO: 2, the molecular weight of the antioxidant peptide modification is 777.89 Da and the isoelectric point is 7.

95.

5. The antioxidant peptide modification according to claim 1, characterized in that: When the amino acid sequence is as shown in SEQ ID NO: 3, the molecular weight of the antioxidant peptide modification is 904.04 Da and the isoelectric point is 5.

84.

6. The antioxidant peptide modification according to claim 1, characterized in that: The preparation method of the modified antioxidant peptide comprises the following steps: synthesizing the complete amino acid sequence of the polypeptide by using a twelve-channel semi-automatic polypeptide synthesizer; desalting and purifying by high performance liquid chromatography reverse phase column chromatography, and determining that the purity is greater than 95%; and determining the amino acid sequence and molecular weight of the synthesized polypeptide by mass spectrometry to obtain the modified antioxidant peptide.

7. Use of the modified marine-derived antioxidant peptide with transmembrane activity and a composition thereof as claimed in any one of claims 1 to 6 in the preparation of a product for scavenging free radicals.

8. The use according to claim 7, characterized in that: The free radical is a 2,2-azino-bis(3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt free radical and / or a hydroxyl free radical.

9. Use of the modified marine-derived antioxidant peptide with transmembrane activity and a composition thereof as claimed in any one of claims 1 to 6 in the preparation of products for resisting ultraviolet radiation and the oxidative damage caused by it.

10. The use according to claim 9, characterized in that: The product for resisting ultraviolet radiation and the oxidative damage caused by it refers to a product for protecting human skin from damage by ultraviolet radiation.

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