Peptides, peptide-containing products, kits and applications

Abstract

This invention provides a polypeptide, a polypeptide product comprising the same, a kit, and applications. The polypeptide, particularly those shown in SEQ ID NO:1 to SEQ ID NO:8, specifically binds to the spike protein of SARS-CoV-2, thereby competitively inhibiting the binding of the SARS-CoV-2 spike protein to hACE2. These polypeptides can competitively reduce or inhibit the binding of the S protein to the ACE2 protein, and therefore can serve as drugs or potential drugs to weaken or inhibit the infectivity of SARS-CoV-2 in humans.

Description

[0001] This application is a divisional application based on the parent application with CN application number 202011447072.2 and application date of December 11, 2020. Technical Field

[0002] This invention relates to the field of peptides, and more specifically, to a peptide, peptide products containing the peptide, kits, and applications thereof. Background Technology

[0003] In order to control or block the spread of the novel coronavirus “SARS-CoV-2” (Severe Acute Respiratory Syndrome Coronavirus 2), there is an urgent need to develop corresponding drugs or preventive agents to curb the spread of the novel coronavirus, especially peptide-based products. Summary of the Invention

[0004] The main objective of this invention is to provide a polypeptide, polypeptide products containing the polypeptide, reagent kits, and applications, offering new means and solutions for curbing the spread of the novel coronavirus.

[0005] To achieve the above objectives, according to one aspect of the present invention, a polypeptide is provided that can specifically bind to the spike protein of SARS-CoV-2, thereby competitively inhibiting the binding of the spike protein to hACE2.

[0006] Furthermore, the peptide can specifically bind to the receptor-binding region of the spike protein of SARS-CoV-2.

[0007] Further, the polypeptide is selected from any one or more of SEQ ID NO: 1 to SEQ ID NO: 8; preferably, the polypeptide is a modified peptide segment; preferably, the modification is a chemical group modification or an amino acid modification; preferably, the chemical group modification is PEG modification; preferably, the PEG modification is a linear PEG modification, a PEG modification with a monofunctional group, or a PEG modification with a bifunctional group; preferably, the PEG modification site is selected from any one or more of the N-terminus, C-terminus, Lys side chain, and Cys side chain of the polypeptide; preferably, the PEG modification is a PEG modification with a molecular weight of 500 to 40000; preferably, the amino acid modification is a hydrophilic amino acid modification or a cysteine ​​modification; preferably, the hydrophilic amino acid modification is... The amino acid modification involves adding 1-4 hydrophilic amino acids to the N-terminus, C-terminus, or NC-terminus of the peptide. More preferably, the hydrophilic amino acids are Glu, Lys, Ser, or Gly. Further preferably, the 1-4 hydrophilic amino acids are selected from any one of the following: Glu-Glu, Lys-Lys, or Ser-Gly-Ser. Preferably, the cysteine ​​modification involves adding a cysteine ​​at any of the following positions of the peptide: N-terminus, C-terminus, NC-terminus, or the middle of the peptide chain. More preferably, adding a cysteine ​​in the middle of the peptide chain includes inserting one or more cysteines in the middle of the peptide chain, or one or more cysteines being linked to the middle of the peptide chain in a branched form.

[0008] According to a second aspect of this application, a polypeptide product is provided, comprising any of the polypeptides described above.

[0009] Furthermore, the peptide product also includes a peptide stabilizer; preferably, the peptide stabilizer includes 150-180 mM NaCl, 100-140 mM polylysine hydrochloride, and water.

[0010] Further, the polypeptide product is a polypeptide drug or polypeptide reagent; preferably, the dosage form of the polypeptide drug is any one of the following: aerosol, solution, pellet, suspension, emulsion, liposome, transdermal preparation, tablet, capsule, pill, powder, granule, suppository, or lyophilized powder for injection; preferably, the route of administration of the polypeptide drug is selected from any one of the following: respiratory administration, mucosal administration, cavity administration, injection administration, or surface administration; preferably, respiratory administration is nasal administration; preferably, cavity administration is selected from rectal, vaginal, or sublingual administration; preferably, injection administration is selected from subcutaneous injection, intravenous injection, intramuscular injection, intraperitoneal injection, intracerebrospinal injection, or infusion.

[0011] Furthermore, the concentration of peptides in the peptide product is 3–10 μM, preferably 5–8 μM.

[0012] According to a third aspect of this application, a kit for detecting SARS-CoV-2 is provided, the kit comprising a detection reagent or a detection chip, the detection reagent comprising any of the aforementioned polypeptides, or the detection chip having any of the aforementioned polypeptides disposed thereon.

[0013] According to a fourth aspect of this application, the use of any of the above-mentioned polypeptides in the preparation of medicaments for the prevention and / or treatment of COVID-19 is provided.

[0014] According to the fifth aspect of this application, the use of any of the above-mentioned peptides in the preparation of a kit for detecting SARS-CoV-2 is provided.

[0015] By applying the technical solution of the present invention, the polypeptides provided in this application, especially the polypeptides shown in SEQ ID NO: 1 to SEQ ID NO: 8, can competitively reduce or inhibit the binding of S protein to ACE2 protein, thereby weakening or inhibiting the infection effect of the novel coronavirus on the human body. Attached Figure Description

[0016] The accompanying drawings, which form part of this application, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. In the drawings:

[0017] Figure 1 A schematic diagram illustrating the specific binding of the polypeptide represented by SEQ ID NO: 7 to the S protein in Example 3 of the present invention is shown; and

[0018] Figure 2 A schematic diagram showing the specific binding of the polypeptide represented by SEQ ID NO: 8 in Example 3 of the present invention to the S protein is shown. Detailed Implementation

[0019] It should be noted that, unless otherwise specified, the embodiments and features described in this application can be combined with each other. The present invention will now be described in detail with reference to the embodiments.

[0020] The novel coronavirus (SARS-CoV-2) causes infection. SARS-CoV-2 belongs to the β-coronavirus genus, is enveloped, and its particles are round or oval, with a diameter of 60–140 nm. It possesses five essential genes, targeting four structural proteins: nucleoprotein (N), viral envelope (E), matrix protein (M), and spike protein (S), as well as an RNA-dependent RNA polymerase (RdRp). The nucleoprotein (N) encapsulates the RNA genome to form the nucleocapsid, which is surrounded by the viral envelope (E). The viral envelope embeds proteins such as the matrix protein (M) and spike protein (S). The spike protein enters cells by binding to angiotensin-converting enzyme 2 (ACE-2). In vitro isolation and culture show that the novel coronavirus can be detected in human respiratory epithelial cells in approximately 96 hours, while isolation and culture in VeroE6 and Huh-7 cell lines takes approximately 4–6 days. Coronaviruses are sensitive to ultraviolet light and heat. They can be effectively inactivated by 56°C for 30 minutes, ether, 75% ethanol, chlorine-containing disinfectants, peracetic acid, and chloroform, while chlorhexidine cannot effectively inactivate the virus.

[0021] In order to develop drugs or detection / diagnostic reagents targeting SARS-CoV-2, this application uses peptide chip technology to screen a batch of peptides that can inhibit the binding of the spike protein to the human receptor protein hACE2, targeting the spike protein that binds to the human receptor protein hACE2. Based on this, the applicant proposes the technical solution of this application.

[0022] A peptide chip is a chip based on a substrate material. The chip includes features with pre-designed quantity, location, and sequence. Each feature is a cluster of peptides with the same sequence. The peptide sequences between features are often different. These features form a high-density peptide array.

[0023] Peptide chip technology is a detection technique based on peptide chips. It utilizes the contact between a variety of peptides on a peptide chip and the sample, then employs fluorescence detection and image acquisition techniques to capture various characteristic signals on the peptide chip (specifically, fluorescence images carrying these characteristic signals). The signal intensity of each feature in the chip is then output, representing the peptide chip detection result data. Based on the sample detection signal output from the peptide chip detection result data, analysis of analytes in samples bound to peptides on the peptide chip, and other sample analysis can be achieved.

[0024] In a first typical embodiment of this application, a polypeptide is provided that can specifically bind to the spike protein of SARS-CoV-2, thereby competitively inhibiting the binding of the spike protein of SARS-CoV-2 to hACE2.

[0025] In a preferred embodiment, the polypeptide is capable of specifically binding to the receptor-binding region of the spike protein of SARS-CoV-2.

[0026] In another preferred embodiment, the polypeptide is able to specifically bind to the hACE2 protein, thereby inhibiting its binding to the spike protein of SARS-CoV-2.

[0027] In a preferred embodiment, the polypeptide is selected from any one or more of SEQ ID NO: 1 to SEQ ID NO: 8. These polypeptides are all capable of specifically binding to the hACE2 protein, thereby competitively inhibiting the binding of the SARS-CoV-2 spike protein to the hACE2 protein.

[0028] To further enhance the affinity of certain peptides, in some preferred embodiments, one or more peptide segments of the aforementioned peptides are modified peptide segments. Specific modifications can be chemical group modifications or amino acid modifications. Preferably, the chemical group modification is PEG modification. More preferably, the PEG modification is linear PEG modification, PEG modification with a monofunctional group, or PEG modification with a difunctional group (the functional group can be -NHS, -OH, -Mal, -NH2, or -COOH; the monofunctional group is selected from any one of these, and the difunctional group is selected from any two of these). Preferably, the PEG modification site is selected from any one or more of the N-terminus, C-terminus, Lys side chain, and Cys side chain of the polypeptide. Preferably, the PEG modification is a PEG modification with a molecular weight of 500 to 40,000. Preferably, the amino acid modification is hydrophilic amino acid modification or cysteine ​​modification. Preferably, the amino acid modification is hydrophilic amino acid modification or cysteine ​​modification. Preferably, the hydrophilic amino acid modification involves adding 1-4 hydrophilic amino acids at the N-terminus, C-terminus, or both N and C-termini. Preferably, the hydrophilic amino acid is Glu, Lys, Ser, or Gly. Preferably, the 1-4 hydrophilic amino acids are selected from any one of the following: Glu-Glu, Lys-Lys, or Ser-Gly-Ser.

[0029] The aforementioned PEG modification or hydrophilic amino acid modification can increase the hydrophilicity of peptides. PEG modification has advantages such as prolonged half-life, reduced toxicity, and enhanced physical, chemical, and biological stability.

[0030] In some embodiments, to better achieve targeted coupling of the peptide, any one or more peptide segments in the aforementioned peptide can be cysteine-modified peptide segments. Specifically, this includes, but is not limited to, adding cysteine ​​at the N-terminus, C-terminus, or NC-terminus of the peptide segment, or adding cysteine ​​in the middle of the peptide chain. When adding cysteine ​​in the middle of the peptide chain, one or more cysteines can be inserted into the middle of the peptide chain (i.e., inserted between two amino acid residues), or one or more cysteines can be linked to the middle of the peptide chain in a branched form (i.e., as a side chain of a certain amino acid in the middle of the peptide chain).

[0031] In a second typical embodiment of this application, a polypeptide product is provided, comprising any of the aforementioned polypeptides. This polypeptide can serve as a polypeptide drug product, capable of reducing or inhibiting the binding of the spike protein to the hACE2 protein by binding to the spike protein of SARS-CoV-2, thereby weakening or blocking SARS-CoV-2 infection in humans. That is, this polypeptide drug product can be used to prevent and / or treat COVID-19. The aforementioned polypeptide can also serve as a detection or diagnostic product, specifically binding to the spike protein of SARS-CoV-2 to determine whether a sample to be tested contains SARS-CoV-2, thus diagnosing whether the sample is a SARS-CoV-2 positive sample.

[0032] To further improve the stability of its active ingredient—the polypeptide—the polypeptide product also includes a polypeptide stabilizer. Any reagent or component that can maintain the storage stability of the polypeptide can be used as the polypeptide stabilizer in this application.

[0033] In a preferred embodiment of this application, the peptide stabilizer comprises 150–180 mM NaCl, 100–140 mM polylysine hydrochloride, and water. More preferably, the peptide stabilizer comprises 153–158 mM NaCl, 110–130 mM polylysine hydrochloride, and water; even more preferably, the peptide stabilizer comprises 154 mM NaCl, 126.4 mM polylysine hydrochloride, and water. Specifically, in the peptide stabilizer, the concentration of NaCl can be 150mM, 151mM, 152mM, 153mM, 154mM, 155mM, 156mM, 157mM, 158mM, 159mM, 160mM, 161mM, 162mM, 163mM, 164mM, 165mM, 166mM, 167mM, 168mM, 169mM, 170mM, 171mM, 172mM, 173mM, 174mM, 175mM, 176mM, 177mM, 178mM, 179mM, or 18... 0 mM; the concentration of polylysine hydrochloride can be 110 mM, 111 mM, 112 mM, 113 mM, 114 mM, 115 mM, 116 mM, 117 mM, 118 mM, 119 mM, 120 mM, 121 mM, 122 mM, 123 mM, 124 mM, 125 mM, 126 mM, 127 mM, 128 mM, 129 mM, 130 mM, 131 mM, 132 mM, 133 mM, 134 mM, 135 mM, 136 mM, 137 mM, 138 mM, 139 mM or 140 mM.

[0034] In a preferred embodiment of this application, the concentration of the polypeptide in the above-mentioned polypeptide product is 3-10 μM, preferably 5-8 μM, and more preferably 5 μM. More specifically, it can be 3 μM, 4 μM, 5 μM, 6 μM, 7 μM, 8 μM, 9 μM, or 10 μM.

[0035] In some preferred embodiments, the above-mentioned polypeptide product is a polypeptide drug, specifically a polypeptide, the various modified polypeptides described above, or their stereoisomers, derivatives, or soluble salts thereof. The specific formulation of this polypeptide drug is not limited, and in this application includes, but is limited to, aerosols, solutions, pellets, suspensions, emulsions, liposomes, transdermal preparations, tablets, capsules, pills, powders, granules, suppositories, or lyophilized powder injections. Depending on the specific dosage form, the above-mentioned polypeptide drug can be administered via a suitable route of administration. In this application, the route of administration is selected from any of the following: respiratory administration, mucosal administration, cavity administration, injection administration, or surface administration. Preferably, respiratory administration is via nasal cavity administration; preferably, cavity administration is selected from rectal, vaginal, or sublingual administration; preferably, injection administration is selected from subcutaneous injection, intravenous injection, intramuscular injection, intraperitoneal injection, intracerebrospinal injection, or infusion.

[0036] The specific dosage of the aforementioned polypeptide drugs can be either a preventative or therapeutic dose. In clinical practice, the doctor determines the dosage based on factors such as the individual's age, weight, and the severity of their condition.

[0037] In a third typical embodiment of this application, a kit for detecting SARS-CoV-2 is also provided. The kit includes a detection reagent or a detection chip. The detection reagent includes any of the aforementioned polypeptides, or the detection chip is provided with any of the aforementioned polypeptides. Here, the polypeptide in the detection reagent or detection chip is a polypeptide capable of specifically binding to the receptor-binding region of the spike protein of SARS-CoV-2. More preferably, it is a polypeptide shown in SEQ ID NO: 1 to SEQ ID NO: 8.

[0038] In a fourth typical embodiment of this application, the use of any of the above-mentioned polypeptides in the preparation of a medicament for treating COVID-19 is also provided. Specific drug dosage forms and administration methods are as described above.

[0039] In a fifth typical embodiment of this application, the application of any of the above-mentioned peptides in the preparation of a kit for detecting SARS-CoV-2 is also provided.

[0040] The beneficial effects of this application will be further illustrated below with reference to specific embodiments.

[0041] Example 1: Screening of peptides

[0042] In this embodiment, candidate peptide drugs with anti-novel coronavirus activity are screened using the receptor-binding domain (RBD, i.e., amino acids 319 to 541 of the S protein) of the spike protein (S) as the target protein. The specific screening steps are as follows:

[0043] 1. Fluorescent labeling of the novel coronavirus S-RBD protein (recombinant protein, specific sequence SEQ ID NO: 9) remember. Specifically, a fluorescent labeling reagent kit (Alexa) was used. Fluorescent labeling was performed using the 555 Protein Labeling Kit (A20174), and the specific procedure is as follows.

[0044] (1) Procedure for S-RBD protein fluorescent labeling experiment:

[0045] a. Prepare a 1M sodium carbonate buffer solution: Add 1 ml of distilled water to the glass bottle containing component B (84 mg of sodium carbonate), and mix well to dissolve.

[0046] b. Diluting the protein: If the protein concentration is greater than 2 mg / mL, the protein needs to be diluted to 2 mg / mL (for example, 1 mg of protein dissolved in 0.5 mL of 1X PBS or 0.1 M sodium carbonate buffer).

[0047] c. Adjust pH: Add 50 μL of 1M sodium carbonate buffer (from step a) to 0.5 mL of 1X PBS containing the protein; the final pH should be 7.5–8.3. This step can be skipped if the protein is already in sodium carbonate buffer (pH 8.0–8.3) or PBS (pH 8.0).

[0048] d. Prepare the labeling reaction mixture: Add the protein solution (from step c) to a glass vial containing the reaction dye (component A). Cap the vial and gently invert or gently blow with a pipette tip to mix thoroughly. Stir the reaction mixture, avoiding foaming.

[0049] e. Incubation of the labeled reaction: Stir magnetically at low speed for 1 hour at room temperature. Take care to avoid foaming of the solution.

[0050] Note: Since preparing the purification column takes about 15 minutes, you can start filling the purification column during the labeling reaction in step e.

[0051] (2) Procedure for purifying labeled S-RBD protein:

[0052] a. Prepare the purification column / purification resin: Assemble the purification column and add purification resin (component C) to 3 cm above the top. Add 1X PBS to check if the buffer flows smoothly. If the buffer flow is obstructed, remove the resin, clean the glass filter, and refill the resin / buffer. Allow excess buffer to drain before adding the labeled protein.

[0053] b. Prepare 1X elution buffer: Dilute 10X elution buffer (component D) 10-fold with distilled water at room temperature. Under normal circumstances, the amount used for a single purification should not exceed 10 mL.

[0054] c. Protein purification: Remove the funnel. Pour the reaction solution onto the purification column resin; wait for the reaction solution to dissolve into the resin. Elute the protein with elution buffer or 1X PBS, collect and retain all residual solution.

[0055] d. Calculate DOL (Delta of Labelling) and store the labeled proteins.

[0056] 2. The fluorescently labeled SARS-CoV-2 S-RBD protein was serially diluted and detected using a peptide microarray. Specific peptides that interact with RBD, and the fluorescence signal of these peptides decreases with increasing dilution.

[0057] The peptide chip detection method is as follows:

[0058] 1) Sample preparation

[0059] The experimental concentrations of the fluorescently labeled S-RBD protein were set at seven concentration gradients (1:500, 1:1000, 1:5000, 1:10000, 1:50000, 1:50000, 1:500000 and 1:5000000 series dilutions) for the test samples to be used.

[0060] 2) Chip hydration and assembly

[0061] The chip was placed in a chip hydration apparatus, and ultrapure water was added to cover the chip. Hydration was carried out on a track shaker at 55±5 rpm / min for 20 minutes. Then, isopropanol was sprayed onto the chip surface, and the chip was centrifuged to dry. The dried chips were then assembled into an assay cassette according to the experimental design.

[0062] 3) Sample and chip incubation combined

[0063] Add the diluted sample at a rate of 90 μL / well to the assembled chip and incubate it on a constant temperature shaker for 1 hour.

[0064] 4) Sample cleaning

[0065] The essay cassette was placed in a plate washer for cleaning.

[0066] 5) Imaging

[0067] The chip in the assay cassette was disassembled, cleaned, dried, and then reassembled into the imaging cassette. It was then placed in Molecular Devices' ImageXpress micro 4 imager for scanning and imaging. Each sample ultimately yielded a TIFF image file, which is the raw data.

[0068] In this embodiment, a peptide technology platform is used to capture signals from target proteins and peptide chip aggregates, and then the signals are converted to obtain data. The instruments and equipment used in the peptide chip technology include peptide chips (e.g., HealthTell V13 chips, model P / N: 600001V13 Slides), fluorescence imagers (e.g., Melecular DeviceImage Xpress Micro-4), chip centrifuges (e.g., Labnet C1303T-230V), plate washers (e.g., BioTek Instruments, 405TSUVS), 96-well plate orbital oscillators (e.g., Thermo Scientific, 88880026), and thermostatic mixers (e.g., Eppendorf Thermomixer C).

[0069] 4. Data Preprocessing

[0070] 1) Extract the fluorescence intensity values ​​of the features and output one GPR5 data file and one corner images file. The GPR5 file contains all the information of a sample and the fluorescence intensity information of all features.

[0071] 2) Extract feature fluorescence intensity information from the GPR5 data files of all samples to generate a raw fluorescence intensity (FG, foreground) data matrix. Then, perform a logarithmic transformation on the data of each sample to obtain an LFG (log-transferred foreground) data matrix, and perform z-score normalization to obtain an NLFG (normalized and log-transferred foreground) data matrix. This step also generates a sample chip information file, which includes information such as the sample array location and the chip number used.

[0072] 5. Quality Control

[0073] The samples and system were quality controlled using Health Tell's built-in quality control methods, and the results showed that they were qualified.

[0074] 6. Screening for polypeptide sequences that can specifically bind to S-RBD protein: Select signal peptides that rank highly in signal intensity at different concentrations after excluding background signals and without supersaturation.

[0075] 6.1) Convert each signal intensity into a log10 value, and then for each concentration of S-RBD protein sample (each sample is tested in three technical replicates), calculate the median of the log10 values ​​of the three technical replicates for each feature.

[0076] 6.2) For each concentration median sample, convert the value to rank (descending; for features with the same value, use method = min(lowest rank in group)) for sorting;

[0077] 6.3) For a peptide, if the rank of a peptide is within the top 100 in concentrations of 70% or higher, the peptide is included as a candidate.

[0078] 6.4) Remove irrelevant samples (i.e., peptides ranked in the top 5000 in the blank control and positive standard) from all candidate peptides;

[0079] 6.5) Sort the peptides according to the average rank and select the top-ranked peptides.

[0080] 7. The candidate polypeptide sequences selected above are as follows:

[0081] SEQ ID NO: 1: YAYEYVFFSE;

[0082] SEQ ID NO: 2: PFFFFEG;

[0083] SEQ ID NO: 3: QVVEVFWLFD;

[0084] SEQ ID NO: 4: NYVFFFEG;

[0085] SEQ ID NO: 5: RLEFLFLFE;

[0086] SEQ ID NO: 6: QYLFFLEG;

[0087] SEQ ID NO: 7: PVFLVFPQRG;

[0088] SEQ ID NO: 8: QSLFLEVFS.

[0089] Example 2 Peptide Validation

[0090] Using BLAST software, all candidate peptides were aligned with the target protein (S protein). A bit score > 14 was selected as the threshold for alignment, indicating a certain degree of similarity between the peptide and the target protein sequence. Based on this result, the peptides were further annotated. The results could be adjusted according to specific needs (e.g., to add one or two sequences that can be aligned with the target protein, these could be selected from the alignment results, even if they are not the highest ranked).

[0091] Based on S protein sequence alignment analysis, the similarity between candidate peptides and S protein sequences is calculated, and based on this, candidate peptide sequences can be classified into different types:

[0092] a) Two peptides that are similar to and match the S protein sequence, that is, the corresponding polypeptides are potential linear epitopes of the S protein, or can explain the scientific basis for binding to the S protein.

[0093] b) Peptides that do not match the S protein sequence but have high signal intensity (the other 6) may be non-linear binding epitopes of the protein or may originate from unknown binding epitopes.

[0094] Example 3

[0095] The validation results of the above polypeptide sequences at known binding sites of the S-RBD protein, and the discovery of potential binding sites.

[0096] Among peptides that bind to the S-RBD protein and rank highly in signal intensity, if these peptides have weak binding affinity to the S-RBD protein or are not specifically bound to the S-RBD protein, their signal intensity will decrease with decreasing S-RBD protein concentration, causing them to rank lower relative to other peptides. Conversely, if the peptide signal intensity does not decrease with decreasing S-RBD protein concentration and remains ranked highly in the peptide sequence, this verifies that the peptide specifically binds to the S-RBD protein and that the binding affinity is strong.

[0097] The SARS-CoV-2 S-RBD protein mediates the entry of the virus into host cells for infection by binding to angiotensin-converting enzyme 2 (ACE2). When the polypeptide binds to the S-RBD protein, the S-RBD protein is unable to bind to the ACE2 receptor or its binding force is weakened, thereby blocking or reducing the virus's infection of host cells, and thus has a competitive inhibitory effect on viral infection.

[0098] The specific design and validation experiments are as follows: Seven dilution concentrations of S-RBD protein were designed (dilution factors: 500, 1000, 5000, 10000, 50000, 500000, and 5000000). A blank control group and a positive standard control group were also designed. The experimental results are shown in Table 1. The candidate peptides mentioned above maintained their high ranking among all peptide signals with increasing concentration dilution, and did not decrease with increasing concentration gradient. The signal ranking of the same peptide in the blank control and standard control groups was significantly different from that of the S-RBD binding signal, indicating a strong and specific binding affinity between these peptides and the S-RBD protein.

[0099] The binding sites of peptides PVFLVFPQRG and QSLFLEVFS to S-RBD are shown in Table 1. Figure 1 and Figure 2 As shown, Figure 1 and Figure 2 The binding sites of the peptides and the S protein are marked in the diagram (the two peptides in the diagram bind to the S protein in different directions, which is why they appear to have different structures). Figure 1 GLN6 indicates that the amino acid at position 6 (indicated by the arrow) is glutamine; Figure 2 PRO60 indicates that the amino acid at position 60 (as indicated by the arrow) is proline.

[0100] Table 1. Ranking of binding strength of candidate peptides at different concentration dilutions at known and potential binding sites of S-RBD protein.

[0101]

[0102] As can be seen from the binding strength ranking in the table above, compared with the control, the binding strength of the above 8 polypeptide sequences to the S protein is stronger.

[0103] In another specific embodiment, the peptides of the present invention were prepared as an aqueous solution as the experimental group, with positive and negative controls set up. These were separately mixed with SARS-CoV-2 pseudovirus and incubated at ambient temperature. After incubation, they were inoculated into pre-prepared cells for further incubation, i.e., a pseudovirus neutralization experiment was performed. The neutralization experiment showed that the peptides of the present invention could all block pseudovirus infection of host cells to a certain extent. Based on the blocking results, the anti-SARS-CoV-2 infection ability of each peptide can be further evaluated.

[0104] As can be seen from the description of the above embodiments, the peptides screened in this application can competitively reduce or inhibit the binding of S protein to ACE2 protein, thereby weakening or inhibiting the infection effect of SARS-CoV-2 on the human body.

[0105] 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. A polypeptide, characterized in that, The polypeptide is capable of specifically binding to the spike protein of SARS-CoV-2, thereby competitively inhibiting the binding of the spike protein to hACE2. The polypeptide is capable of specifically binding to the receptor binding domain of the spike protein of SARS-CoV-2. The polypeptide is the polypeptide shown in SEQ ID NO:

8.

2. A polypeptide composition, characterized in that, The polypeptide composition comprises the polypeptide shown in SEQ ID NO: 8, and further comprises any one or more of the polypeptides shown in SEQ ID NO: 2 to SEQ ID NO:

6.

3. A polypeptide product, characterized in that, The polypeptide product comprises the polypeptide of claim 1 or the polypeptide composition of claim 2.

4. The polypeptide product of claim 3, wherein, The polypeptide product further comprises a polypeptide stabilizer.

5. The polypeptide product of claim 4, wherein, The polypeptide stabilizer comprises 150-180 mM NaCl, 100-140 mM polylysine hydrochloride and water.

6. The polypeptide product of claim 4, wherein, The polypeptide product is a polypeptide drug or a polypeptide reagent.

7. The polypeptide product of claim 6, wherein, The polypeptide drug is in any one of the following dosage forms: aerosol, solution, suspension, emulsion, liposome, transdermal, tablet, capsule, pill, powder, granule or suppository.

8. The polypeptide product of claim 7, wherein, The powder comprises a freeze-dried powder injection, and the pill comprises a drop pill.

9. The polypeptide product of claim 6, wherein The polypeptide drug is administered in any one of the following ways: cavity administration, injection administration or surface administration.

10. The polypeptide product of claim 9, wherein the polypeptide product is a protein. The cavity administration comprises respiratory tract administration, and the surface administration comprises mucosal administration.

11. The polypeptide product of claim 10, wherein The respiratory tract administration is nasal cavity administration.

12. The polypeptide product of claim 9, wherein The cavity administration is selected from rectal, vaginal or sublingual administration.

13. The polypeptide product of claim 9, wherein The injection administration is selected from subcutaneous injection, intravenous injection, intramuscular injection, intraperitoneal injection, intracisternal injection or infusion.

14. The polypeptide product of claim 6, wherein The concentration of the polypeptide in the polypeptide product is 3-10 μM.

15. The polypeptide product of claim 14, wherein, The concentration of the polypeptide in the polypeptide product is 5-8 μM.

16. A kit for detecting SARS-CoV-2, characterized in that, The kit comprises a detection reagent or a detection chip, wherein the detection reagent comprises the polypeptide of claim 1 or the polypeptide composition of claim 2, or the detection chip is provided with the polypeptide of claim 1 or the polypeptide composition of claim 2.

17. Use of the polypeptide of claim 1 or the polypeptide composition of claim 2 in the preparation of a drug for preventing and / or treating COVID-19.

18. Use of the polypeptide of claim 1 or the polypeptide composition of claim 2 in the preparation of a kit for detecting SARS-CoV-2.

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