Polypeptide targeting novel coronavirus COVID-19 and application of polypeptide
A technology for COVID-19 and coronavirus, applied in the field of peptides targeting the new coronavirus COVID-19, can solve the difficult problems of effective treatment of coronavirus, and achieve the effect of enriching sequence diversity, ensuring efficiency and small molecular weight
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Embodiment 1
[0091] Example 1 Biotin or FITC labeling of S protein
[0092] In this example, the S protein (SEQ ID NO: 29) is first labeled with biotin or FITC, and the steps are as follows:
[0093] 0.15 mol / L NaCl solution and 0.15 mol / L NaHCO at pH=9.0 3 -Na2 CO 3 The buffer solution was mixed at a molar ratio of 9:1, and protein S was added to the mixture for protein dissolution, so that the final concentration of protein S was 10 mg / mL;
[0094] Add biotin or FITC to the S protein solution, place in an ice bath, and magnetically stir overnight in the dark;
[0095] On the second day, the mixed solution was placed in a dialysis bag with a molecular weight cut-off of 200, dialyzed in deionized water for two days, and the water was changed several times;
[0096] Finally, the dialyzed solution was collected and freeze-dried by a freeze dryer to obtain biotin- or FITC-labeled S protein.
Embodiment 2
[0097] Embodiment 2 Establishment of OBOC polypeptide library
[0098] In this example, the OBOC method is used to synthesize a peptide library on resin beads (beads), and the steps are as follows:
[0099] (1) Take 1.8 g of resin beads and put them in the synthesis tube, add a certain amount of DMF to soak the resin for resin swelling, and soak for more than 2 hours;
[0100] (2) Add Fmoc-protected methionine (M) to the synthesis tube, and add a mixture of HOBT (1-hydroxybenzotriazole) and DIC (N,N'-diisopropylcarbodiimide) solution, reacted on a shaking table for more than 2 h; alternately rinsed with methanol and DMF three times, repeatedly blowing up the beads with a straw during the rinse process; then tested the beads in a ninhydrin boiling water bath, and the beads were colorless;
[0101] (3) Add a deprotection agent (20% hexahydropyridine + 80% DMF) for Fmoc deprotection for 10 min, alternately rinse with methanol and DMF three times, and repeatedly blow up the beads...
Embodiment 3
[0108] Example 3 Screening of polypeptides targeting S protein
[0109] The FITC-labeled S protein and the peptide library were incubated in PBS buffer for 2 hours, the luminescence was observed under a fluorescence microscope (excitation wavelength 405 nm), and the observed peptide resins with obvious green fluorescence were selected.
[0110] like figure 1 Shown is the fluorescent microscope picture of FITC-labeled S protein interacting with OBOC polypeptide library. The beads bound to S protein can be observed with green fluorescence, and the peptides that can bind to S protein are selected according to the fluorescence.
[0111] The selected polypeptide resin was washed several times with ultrapure water and dried, using 30% CNBr 3 The ethanol solution was lysed overnight, the supernatant was obtained by centrifugation, and the peptide sequencing was carried out after desalting treatment.
[0112] Exemplarily, Figure 2(A), Figure 2(B), Figure 2(C) and Figure 2(D) are the...
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