Horseshoe crab anti-endotoxin factor analogue peptide molecule, synthetic method and use thereof
An anti-endotoxin and peptide-mimicking technology, applied in the fields of peptide preparation, peptide/protein components, chemical instruments and methods, etc., can solve the problems of high cost, long period of genetic engineering, and unsuitable for human treatment.
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Embodiment 1
[0093] This example relates to the synthesis of S0-040420, which is used to illustrate a manual synthesis method and purification method of the limulus anti-endotoxin factor mimetic peptide of the present invention.
[0094] 1. Synthesis of S0-040420 Peptide Resin
[0095] The peptide sequence of S0 is: RINPTVKRLKWKYKGKFWCP.
[0096] Weigh 2.420g (0.5mmol) of Fmoc-Pro-2-Cl-Trt Resin-040410 and pour it into a synthesis column, add 35mL of DCM-DMF (1:2, v / v) to swell. After 30 min, the solvent was extracted, and the Fmoc protecting group of the amino acid resin was removed with 15 mL of 30% hexahydropyridine in DMF, and after 30 min, it was washed with DCM, MeOH, and DMF in sequence. Weigh 1.176g Fmoc-Cys(Trt)-OH into a 100mL Erlenmeyer flask, add 10mL DMF to dissolve, add 0.780g TBTU, 0.301g HOBt and 35mL 25% DMF solution of diisopropylethylamine in batches. Ninhydrin detection reaction process. After the coupling is completed, remove the reaction solution, wash th...
Embodiment 2
[0111] This example relates to the synthesis of S4-040406 to illustrate another synthesis and purification method of the limulus anti-endotoxin factor mimetic peptide of the present invention.
[0112] 1. Synthesis of S4-040406 peptide resin
[0113] The peptide sequence of S4 is: RLNPTIKRIRWKYKGKFW.
[0114] Weigh 1.35g (0.5mmol) Fmoc-Trp(Boc)-wang Resin-040402 and pour it into a synthesis column, add 25mL DMF to swell. After 30 min, the solvent was extracted, and the Fmoc protecting group of the amino acid resin was removed with 15 mL of 25% hexahydropyridine in DMF, and after 10 min, it was washed with DCM, MeOH, DMF, DCM, and DMF in sequence. Weigh 0.753g Fmoc-Phe-OH into a 100mL Erlenmeyer flask, add 10mL DMF to dissolve, add 0.50mL DIC, 0.301g HOBt and 35mL 25% DMF solution of diisopropylethylamine in batches. Ninhydrin detection reaction process. After the coupling is completed, remove the reaction solution, wash the peptide resin with DCM, MeOH, DMF, DCM, and DMF in s...
Embodiment 3
[0129] This example relates to the synthesis of S1-040420 and S6-040420, which is used to illustrate the synthesis method of the limulus anti-endotoxin factor mimetic peptide of the present invention using a polypeptide synthesizer.
[0130] The peptide sequence of S1 is: RLNPTIKRLKFKYKGKWWCP.
[0131] The peptide sequence of S6 is: PTIKRVKWKYKGKFWCP.
[0132] The 12-channel polypeptide synthesizer Symphony(R) produced by Protein Technologies, Inc. of the United States is used for synthesis, and 12 polypeptides can be synthesized simultaneously in one cycle. Generally, the amount of protected amino acid is 6 times of the scale to be synthesized, and the ratio of activator to amino acid is 1:1. The solvent for swelling, washing and reaction is mainly DMF; the coupling time of each amino acid needs to be designed, which varies greatly; the choice of activator is very important and varies with the amino acid, peptide sequence and peptide chain length.
[0133] Fmoc-AA-...
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