Polypeptide with uric acid-lowering activity and preparation method and application thereof

Abstract

The application discloses a polypeptide with uric acid reducing activity and a preparation method and application thereof, relates to the field of bioactive peptides, and provides a preparation method of the polypeptide with uric acid reducing activity, which takes whey protein as raw material, and obtains the polypeptide with high activity through directional enzymolysis and ultrafiltration purification, and the polypeptide can reduce blood uric acid level by inhibiting xanthine oxidase activity, and the method has the advantages of high efficiency, low cost and easy implementation, can be implemented on a large scale, and is helpful to improving the utilization value of whey protein in the industry, and provides a new way for the development of functional natural anti-uric acid level abnormality or xanthine oxidase activity abnormality related disease products.

Description

Technical Field

[0001] This invention relates to the field of bioactive peptides, and more specifically, to polypeptides with uric acid-lowering activity, their preparation methods, and applications. Background Technology

[0002] Gout is an inflammatory joint disease caused by purine metabolism disorders or impaired uric acid excretion. Its core pathological feature is persistently elevated serum uric acid (SUA) levels, leading to the deposition of monosodium urate (MSU) crystals in the joints and surrounding tissues. This triggers acute inflammatory responses (manifested as severe pain, swelling, and functional impairment) and chronic joint damage. Epidemiological data shows a significant upward trend in the global prevalence of gout, with approximately 55.8 million patients in 2020 and an annual incidence rate of about 0.3%. The incidence rate in men is 3.26 times that in women, and the risk increases with age.

[0003] Currently, clinical treatment mainly relies on xanthine oxidase (XO) inhibitors (such as allopurinol and febuxostat) to reduce uric acid production, or on uricosuric drugs (such as benzbromarone) to accelerate uric acid excretion. However, synthetic XO inhibitors have significant limitations: (1) long-term use can easily cause liver and kidney toxicity and allergic reactions; (2) some drugs (such as febuxostat) may increase the risk of cardiovascular events, limiting their applicable population. In addition, uricosuric drugs are contraindicated in patients with kidney stones, and their efficacy is greatly affected by renal function.

[0004] To address the aforementioned issues, developing safe and effective natural XO inhibitors has become an important direction in drug development. Studies have shown that bioactive peptides derived from various proteins, including dairy products, marine organisms, grains, and legumes, can lower blood uric acid levels by directly inhibiting xanthine oxidase activity or promoting uric acid excretion. However, existing screening methods for XO inhibitory peptides suffer from low targeting specificity and unclear mechanisms, leading to insufficient conversion efficiency of related products.

[0005] In view of this, the present invention is proposed. Summary of the Invention

[0006] The purpose of this invention is to provide a polypeptide with uric acid-lowering activity, its preparation method, and its application.

[0007] This invention is implemented as follows:

[0008] In a first aspect, embodiments of the present invention provide a polypeptide, wherein the polypeptide is selected from any one of a first polypeptide and a second polypeptide having at least 80% identity with the amino acid sequences shown in SEQ ID NO:1~2.

[0009] Secondly, embodiments of the present invention provide a composition comprising the polypeptide described in the foregoing embodiments.

[0010] Thirdly, embodiments of the present invention provide a method for preparing a polypeptide with uric acid-lowering activity, which includes the following steps: hydrolyzing a whey protein solution with a protease and performing solid-liquid separation, and taking the supernatant; wherein the protease includes any one or more of alkaline protease, trypsin, and chymotrypsin;

[0011] The polypeptide with uric acid-lowering activity contains the polypeptide described in the foregoing examples or the composition described in the foregoing examples.

[0012] Fourthly, embodiments of the present invention provide a polypeptide with uric acid-lowering activity, which is prepared by the preparation method described in the foregoing embodiments.

[0013] Fifthly, embodiments of the present invention provide the use of the polypeptide or polypeptide-containing composition as described in the foregoing embodiments, the third polypeptide with an amino acid sequence as shown in SEQ ID NO:3, the preparation method described in the foregoing embodiments, or the polypeptide with uric acid-lowering activity described in the foregoing embodiments in the preparation of a composition with uric acid-lowering activity or xanthine oxidase-inhibiting activity.

[0014] Sixthly, embodiments of the present invention provide the use of the polypeptides described in the foregoing embodiments, the compositions described in the foregoing embodiments, the third polypeptide with an amino acid sequence as shown in SEQ ID NO:3, the preparation method described in the foregoing embodiments, or the polypeptides with uric acid-lowering activity described in the foregoing embodiments in the preparation of medicaments for the prevention, improvement, or treatment of diseases related to abnormal uric acid levels or abnormal xanthine oxidase activity.

[0015] The present invention has the following beneficial effects:

[0016] This invention provides a polypeptide with uric acid-lowering activity and its preparation method. The method uses whey protein as raw material and obtains a highly active polypeptide through targeted enzymatic hydrolysis and ultrafiltration purification. This polypeptide can lower blood uric acid levels by inhibiting xanthine oxidase activity. This method has advantages such as high efficiency, low cost, and ease of implementation. It can be implemented on a large scale and helps to improve the industrial utilization value of whey protein. It provides a new approach for the development of functional natural anti-uric acid level or xanthine oxidase activity-related disease products. Attached Figure Description

[0017] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.

[0018] Figure 1 This is a process flow diagram for the preparation of the XO inhibitory peptide of this application;

[0019] Figure 2 This is a graph showing the degree of hydrolysis of multiple enzymes.

[0020] Figure 3 Graph showing the xanthine oxidase inhibition assay of multi-enzyme hydrolysates;

[0021] Figure 4 The graph shows the xanthine oxidase inhibition rate of alkaline protease hydrolysates under different temperature conditions.

[0022] Figure 5 A graph showing the xanthine oxidase inhibition rate of alkaline protease hydrolysates under different pH conditions;

[0023] Figure 6 Xanthine oxidase inhibition rates for unseparated fractions, <3 kDa, 3–10 kDa, and >3 kDa;

[0024] Figure 7 The total ion chromatogram is LC-MS / MS for MW < 3 kDa;

[0025] Figure 8 This is a diagram showing the docking of peptides KIPAVF, SDISLL, and ASDISLL with xanthine oxidase molecules. Detailed Implementation

[0026] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.

[0027] Membrane separation technology can only separate molecules based on size / charge, unlike column chromatography (such as affinity chromatography) which can achieve specific binding and has limited retention precision. Based on research into whey protein solutions and membrane separation technology, and through extensive experimental verification, the inventors of this application have established a method for separating peptides with XO inhibitory activity from whey protein by synergistically optimizing enzymatic hydrolysis and ultrafiltration parameters. This method significantly improves the enrichment efficiency of target peptides, enhances the utilization efficiency of whey protein, and enables large-scale, low-cost, and high-throughput screening of peptides with XO inhibitory activity.

[0028] Definition of noun

[0029] The terms “first,” “second,” “third,” etc., used in this article are for distinguishing descriptions only and should not be interpreted as indicating or implying relative importance.

[0030] In this invention, the term "identity" percentage refers to the degree to which the amino acids of two polypeptides are identical at equivalent positions when two sequences are optimally aligned. The amino acid sequence identity percentage alignment can be performed using various methods within the art, such as software well-known in the field, including BLAST, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, or CLUSTAL OMEGA.

[0031] The term "ultrafiltration" in this article refers to a membrane separation technology based on molecular weight cutoff (MWCO), typically in the range of 1,000 to 1,000,000 Da, which works through a combination of sieving effect and charge repulsion.

[0032] The term “treatment” in this article includes preventing or alleviating a condition, slowing the onset or development of a condition, reducing the risk of developing a condition, preventing or delaying the development of symptoms associated with a condition, reducing or stopping symptoms associated with a condition, producing a complete or partial reversal of a condition, curing a condition, or a combination of the above.

[0033] In this article, the term "prevention" refers to preventing the onset, development, or worsening of a disease or health problem before or at an early stage, through proactive interventions. Its core objective is to reduce the probability of disease occurrence, delay the onset time, and / or reduce the negative impact of disease on an individual.

[0034] Specific Plan

[0035] On one hand, embodiments of the present invention provide a polypeptide, wherein the polypeptide is selected from either a first polypeptide (SDISLL) or a second polypeptide (ASDISLL) whose amino acid sequences are at least 80% identical to the sequences shown in SEQ ID NO: 1~2.

[0036] In an optional embodiment, the at least 80% means: 80%, 82%, 84%, 86%, 88%, 90%, 92%, 94%, 96%, 98%, 99% or more, or 100%.

[0037] On the other hand, embodiments of the present invention provide a composition comprising the polypeptide described in any of the foregoing embodiments.

[0038] In an optional embodiment, the composition comprises at least two of a first polypeptide, a second polypeptide, and a third polypeptide (KIPAVF) having at least 80% identity with the sequence shown in SEQ ID NO:3.

[0039] In an optional embodiment, the at least 80% means: 80%, 82%, 84%, 86%, 88%, 90%, 92%, 94%, 96%, 98%, 99% or more, or 100%.

[0040] In an optional embodiment, the composition includes any one of the following: (I) a combination of a first polypeptide and a second polypeptide; (II) a combination of a second polypeptide and a third polypeptide; (III) a combination of a first polypeptide and a third polypeptide; or a combination of a first polypeptide, a second polypeptide, and a third polypeptide.

[0041] On the other hand, embodiments of the present invention provide a method for preparing a polypeptide or xanthine oxidase inhibitory peptide with uric acid-lowering activity, which includes the following steps: hydrolyzing a whey protein solution with a protease and performing solid-liquid separation, and taking the supernatant; the protease includes any one or more of alkaline protease, trypsin, chymotrypsin and pepsin.

[0042] In an optional embodiment, the hydrolysis using alkaline protease is performed at a temperature of 50–60°C, with the pH maintained at 8.5–9.5, for a time of 60–360 min. Specifically, the temperature can be any one or a range between any two of 50, 52, 54, 56, 58, and 60°C; and the pH can be any one or a range between any two of 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, and 9.5.

[0043] In an optional embodiment, the hydrolysis using trypsin is carried out at a temperature of 30-45°C, with the pH maintained at 7.5-8.5, for a time of 60-360 min; specifically, the temperature can be any one or any two of 30, 32, 34, 36, 37, 38, 40, 42, 44, and 45°C; specifically, the pH can be any one or any two of 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, and 8.5.

[0044] In an optional embodiment, the hydrolysis using chymotrypsin is performed at a temperature of 30–45°C, with the pH maintained at 7.5–8.5, for a time of 60–360 min. Specifically, the temperature can be any one or any two of the following: 30, 32, 34, 36, 37, 38, 40, 42, 44, and 45°C; and the pH can be any one or any two of the following: 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, and 8.5.

[0045] In an optional embodiment, the hydrolysis using pepsin is performed at a temperature of 30–45°C, with the pH maintained at 1.5–2.5, for a time of 60–360 min. Specifically, the temperature can be any one or any two of the following: 30, 32, 34, 36, 37, 38, 40, 42, 44, and 45°C; and the pH can be any one or any two of the following: 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, and 2.5.

[0046] In an optional embodiment, the preparation method further includes: ultrafiltration of the supernatant to obtain a product with a molecular weight <3kDa.

[0047] In an optional embodiment, the preparation method further includes, prior to hydrolysis, adjusting the temperature and pH of the whey protein solution to the temperature and pH suitable for the protease.

[0048] In an optional embodiment, the time of 60 to 360 min can specifically be any one or any two of the following: 60, 80, 100, 120, 140, 160, 180, 200, 220, 240, 260, 280, 300, 320, 340, and 360 min.

[0049] In an optional embodiment, the amount of protease added (the ratio of protease activity to whey protein content) is 4000~6000 U / g, specifically any one or any two of 4000, 4250, 4500, 4750, 5000, 5250, 5500, 5750, and 6000 U / g.

[0050] In an optional embodiment, the whey protein solution contains whey protein at a mass percentage of 1% to 10%, specifically within any one or any two of the following: 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, and 10%.

[0051] In an optional embodiment, the whey protein solution is prepared using deionized water.

[0052] In an optional embodiment, after hydrolyzing the whey protein solution with a protease and before solid-liquid separation, the preparation method further includes a step of inactivating the protease in the hydrolysis product. The enzyme can be inactivated by heat.

[0053] In an optional embodiment, the thermal deactivation includes heating the hydrolysate at 80-110°C for 1-30 minutes. Specifically, the heating temperature can be any one or any two of 80, 85, 90, 95, 100, 105, and 110°C. The heating time can be any one or any two of 1, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, and 30 minutes.

[0054] In an optional embodiment, the solid-liquid separation method includes centrifugation.

[0055] In an optional embodiment, the centrifugation conditions include: 0~10℃, 6000~10000×g, 5~30min. Specifically, the temperature can be any one or any two of 0, 2, 4, 6, 8, and 10℃. The centrifugation acceleration can be any one or any two of 6000, 7000, 8000, 9000, and 10000×g. The temperature can be any one or any two of 5, 10, 15, 20, 25, and 30min.

[0056] On the other hand, embodiments of the present invention provide polypeptides or xanthine oxidase inhibitory peptides with uric acid-lowering activity, which are prepared by the preparation method described in any of the foregoing embodiments.

[0057] On the other hand, embodiments of the present invention provide the use of the first polypeptide, second polypeptide, third polypeptide or composition as described in any of the foregoing embodiments, the preparation method described in any of the foregoing embodiments, or the polypeptide or xanthine oxidase inhibitory peptide with uric acid-lowering activity described in any of the foregoing embodiments in the preparation of compositions with uric acid-lowering activity or xanthine oxidase inhibitory activity.

[0058] Furthermore, embodiments of the present invention also provide the use of the first polypeptide, second polypeptide, third polypeptide or composition as described in any of the foregoing embodiments, the preparation method described in any of the foregoing embodiments, or the polypeptide or xanthine oxidase inhibitory peptide with uric acid-lowering activity described in any of the foregoing embodiments in medicaments for the prevention, improvement or treatment of diseases related to abnormal uric acid levels or abnormal xanthine oxidase activity.

[0059] In an optional embodiment, the diseases associated with abnormal uric acid levels or abnormal xanthine oxidase activity include at least one of gout, hyperuricemia, and uric acid nephropathy.

[0060] The features and performance of the present invention will be further described in detail below with reference to embodiments.

[0061] Example 1

[0062] A method for preparing a polypeptide, comprising the following steps.

[0063] (1) Hydrolyzed whey protein solution

[0064] A 5% (w / w) whey protein solution was prepared using deionized water. The temperature and pH of the whey protein solution were adjusted to the optimal hydrolysis temperature and pH for protease adaptation. After the temperature and pH stabilized, protease was added to the whey protein solution at a ratio of 4800 U / g of protease activity to whey protein mass for hydrolysis. An alkaline protease was used, with an optimal hydrolysis temperature of 55℃ and a pH of 9.0. The temperature was kept constant during hydrolysis, and the pH of the hydrolysis system was kept stable using 1 mol / L NaOH. Samples were taken at different time points (0, 10, 20, 30, 40, 50, 60, 90, 120, 150, 180, 210, 240, 300, and 360 min) during the hydrolysis process.

[0065] (2) The hydrolysis products were subjected to enzyme inactivation and centrifugation.

[0066] The extracted hydrolysate sample was quickly boiled in a 100℃ water bath for 20 min to inactivate the enzyme. After inactivation, the sample was cooled and centrifuged at 8000×g for 20 min at 4℃. The supernatant was then stored at -20℃ for subsequent analysis.

[0067] (3) Ultrafiltration

[0068] The obtained supernatant was filtered using an ultrafiltration membrane with a molecular weight cutoff of 3 kDa to obtain components with a molecular weight cutoff of <3 kDa.

[0069] The process flow diagram can be referred to. Figure 1 .

[0070] Example 2

[0071] A method for preparing a polypeptide is generally the same as in Example 1, except that trypsin is used for hydrolysis, and the optimal hydrolysis temperature is 37°C and the pH is 8.0.

[0072] Example 3

[0073] A method for preparing a polypeptide is generally the same as in Example 1, except that: chymotrypsin is used for hydrolysis, and the optimal hydrolysis temperature is 37°C and the pH is 8.0.

[0074] Example 4

[0075] A method for preparing a polypeptide is generally the same as in Example 1, except that: pepsin is used for hydrolysis, and the optimal hydrolysis temperature is 37°C and the pH is 2.0.

[0076] Example 5

[0077] The peptides were prepared using the methods described in Examples 1-4, and the degree of hydrolysis and xanthine oxidase inhibition rate of the hydrolysis product from step (1) were tested.

[0078] (A) Degree of hydrolysis

[0079] The degree of hydrolysis (DH) of the hydrolysate was determined using the o-phthalaldehyde (OPA) method. The sample was diluted 400-fold. 3 mL of the diluted sample (with 3 mL of deionized water as a blank) was mixed with 3 mL of OPA reagent and reacted at room temperature in the dark for 5 min. The absorbance was measured at 340 nm using a UV spectrophotometer. The degree of hydrolysis (DH) was calculated (%) based on the leucine standard curve and formula. In the formula: Leucine content (μg / mL) of the hydrolysis products calculated based on the standard curve; : Leucine content in the sample before hydrolysis calculated according to the standard curve (μg / mL); N: Sample dilution factor, which is 400 in this example; 131.17: Molar mass of leucine (g / mol); X: Initial protein concentration (g / L). The total peptide bond content of the protein is 8.8 mmol / g per gram of whey protein.

[0080] Hydrolysis curves are shown Figure 2 For whey protein, alkaline protease exhibited the strongest hydrolytic ability, rapidly increasing its degree of hydrolysis to approximately 12% within the first 60 minutes, after which the hydrolysis rate gradually slowed, eventually reaching 16.84 ± 0.39%. Trypsin showed slightly weaker hydrolytic ability than alkaline protease, with its degree of hydrolysis rapidly increasing within the first 60 minutes, followed by a gradual slowdown, ultimately reaching 8.87 ± 0.09%. The degrees of hydrolysis for chymotrypsin and pepsin were 7.86 ± 0.24% and 6.27 ± 0.10%, respectively.

[0081] (B) Xanthine oxidase inhibition rate

[0082] The hydrolysate from step (1) was diluted 10-fold with 0.2 mol / L PBS solution (pH=7.5) for subsequent assays. 75 μL of the diluted sample solution and 75 μL of 0.05 U / mL xanthine oxidase (XO) solution were added to a 96-well plate and mixed thoroughly. The plate was incubated at 37°C for 15 min. Subsequently, 75 μL of 0.7 mmol / L xanthine (XA) substrate solution was added, and the reaction was continued at room temperature for 20 min. The absorbance was measured at 295 nm using a microplate reader. All experiments were repeated three times. The XO inhibition rate was calculated according to the following formula. The xanthine oxidase inhibitory activity of the hydrolysate at different dilutions was measured, and the results were calculated. value. In the formula: : Absorbance after adding hydrolyzed sample solution; Use PBS buffer solution instead of PBS buffer solution to measure the absorbance of the hydrolyzed sample; : Absorbance of PBS buffer solution instead of XO solution; Use PBS buffer solution instead of the absorbance of hydrolyzed sample and XO solution.

[0083] Results of xanthine oxidase inhibition rate are shown in Figure 3 During the hydrolysis of whey protein, the xanthine oxidase inhibition rate of the alkaline protease hydrolysate was the highest. With increasing hydrolysis time, the inhibition rate first increased and then decreased, reaching its peak at 240 min (70.59 ± 0.97%). The xanthine oxidase inhibition rate of trypsin hydrolysate of whey protein steadily increased in the early stages of hydrolysis, reaching its highest value of 60.43 ± 0.62% at 240 min. The inhibition rate of chymotrypsin hydrolysate of whey protein showed a similar trend to that of trypsin, with a maximum inhibition rate of 54.61 ± 1.08%.

[0084] Example 6

[0085] To verify the effect of different hydrolysis temperatures on the inhibition rate of xanthine oxidase in the product.

[0086] Using the preparation method provided in Example 1, multiple experimental groups were set up with hydrolysis temperature as a single-factor variable. Hydrolysis was carried out at 40, 45, 50, 55, and 60℃, respectively. After hydrolysis for 240 min, the xanthine oxidase inhibition rate of the hydrolysis product was tested. The test method is described in Example 5. The results are shown below. Figure 4 .

[0087] The results show that the optimal hydrolysis temperature for alkaline protease is 55℃.

[0088] Example 7

[0089] To verify the effect of different hydrolysis pH on the inhibition rate of xanthine oxidase in the product.

[0090] Using the preparation method provided in Example 1, multiple experimental groups were set up with hydrolysis pH as a single-factor variable. Hydrolysis was performed at pH values ​​of 7.5, 8.0, 8.5, 9.0, and 9.5, respectively. After hydrolysis for 240 min, the xanthine oxidase inhibition rate of the hydrolysis products was tested. The test method is described in Example 5. Results are shown below. Figure 5 .

[0091] The results show that the optimal pH for the hydrolysis of alkaline protease is 9.

[0092] Example 7

[0093] The effect of ultrafiltration membrane on the inhibition rate of xanthine oxidase in the product was verified.

[0094] Using the preparation method provided in Example 1, after hydrolysis for 240 min, multiple experimental groups were set up with ultrafiltration as a single-factor variable to obtain fractions <3 kDa, 3~10 kDa, and >3 kDa, respectively. The xanthine oxidase inhibition rate of the products was tested, and the test method is described in Example 5. The results are shown below. Figure 6 .

[0095] The results show that the xanthine oxidase inhibition rate was highest in the <3 kDa fraction, reaching 72.21 ± 1.25%.

[0096] Example 8

[0097] Mass spectrometry was used to identify the components with <3kDa after hydrolysis for 240 min in Example 1. The results are shown in [Figure 1]. Figure 7 The amino acid sequences selected to determine the optimal peptides are: KIPAVF (SEQ ID NO:3), SDISLL (SEQ ID NO:1), and ASDISLL (SEQ ID NO:2).

[0098] The xanthine oxidase inhibitory activities of KIPAVF, SDISLL, and ASDISLL were tested, and the test methods are described in Example 5. The values ​​were 9.24 mM, 6.78 mM, and 13.01 mM, respectively.

[0099] The identified peptide and xanthine oxidase were molecularly docked, and the crystal structure of xanthine oxidase was obtained from the PDB database (PDB ID: 1FO4). Molecular docking was performed using MOE to validate the model predictions, and intermolecular interactions were visualized using PyMOL. The results are shown in [link to results]. Figure 8 The inhibition mode of the selected peptide on xanthine oxidase was determined.

[0100] The results showed that all peptides could form stable binding with xanthine oxidase, and multiple hydrogen bonds were formed near the active site. KIPAVF formed four hydrogen bonds with xanthine oxidase, two of which acted on the LYS-771 residue, and the other two bound to PRO-1076 and ASN-1073, respectively. SDISLL and ASDISLL formed two and five hydrogen bonds, respectively. ASDISLL had the most hydrogen bonds of all peptides, forming two independent hydrogen bonds with LYS-771, and simultaneously interacting with multiple residues such as GLU-802, SER-876, and LEU-648, forming a stable network structure. The above docking results analysis shows that these highly active peptides can bind well to the active site of xanthine oxidase in terms of spatial structure and form stable hydrogen bonds with a variety of key residues, especially LYS-771, GLU-802, and ASN-1073 residues. This indicates that they play an important role in the binding process between peptides and enzymes and may be the core targets for peptides to inhibit xanthine oxidase activity.

[0101] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. The use of a polypeptide or a composition containing the polypeptide with an amino acid sequence as shown in SEQ ID NO:1 or 2 in the preparation of a composition having uric acid-lowering activity.

2. The use of a polypeptide or a composition containing the polypeptide as shown in SEQ ID NO:1 or 2 in the preparation of a medicament for the prevention, improvement or treatment of diseases related to abnormal uric acid levels or abnormal xanthine oxidase activity; wherein the abnormal uric acid levels or abnormal xanthine oxidase activity are at least one of gout, hyperuricemia and uric acid nephropathy.

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