Hydrophis cyanocinctus antibacterial peptide modified body HC14 and its preparation method and application
A technology of HC14 and antimicrobial peptides, applied in the field of biomedicine, can solve problems such as antibiotic resistance of pathogenic microorganisms and threats to human health, and achieve the effects of low hemolytic activity, broad-spectrum and high-efficiency antibacterial activity, and small molecular weight
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Embodiment 1
[0014] Chemical Synthesis of Antimicrobial Peptide HC14 from Sea Snake Modified Body
[0015] Qinghuan Sea Snake Antimicrobial Peptide QHA is a linear polypeptide encoded by a gene, containing 30 amino acid residues, with a molecular weight of 3628.5Da and an isoelectric point of 12.61. The complete sequence of Qinghuan sea snake antimicrobial peptide QHA is: Lys 1 Phe 2 Phe 3 Lys 4 Arg 5 Leu 6 Leu 7 Lys 8 Ser 9 Val 10 Arg 11 Arg 12 Ala 13 Val 14 Lys 15 Lys 16 Phe 17 Arg 18 Lys 19 Lys 20 Pro 21 Arg 22 Leu 23 Ile 24 Gly 25 Leu 26 Ser 27 Thr 28 Leu 29 Leu 30 . According to the amino acid sequence of Qinghuan sea snake antibacterial peptide QHA, the modified HC14 was designed and obtained by molecular transformation method, and it was chemically synthesized by peptide solid-phase synthesis method. The specific preparation method is as follows:
[0016] I. Preparation method of HC14: According to the above amino acid sequence of HC14, its full sequenc...
Embodiment 2
[0022] HC14 Pharmacological Experiment:
[0023] 1. Determination of antibacterial activity of HC14:
[0024] (1) Pick the test strains stored on the slope and evenly spread them on the MH solid medium (purchased from Qingdao Haibo Biotechnology Co., Ltd.) plate, and place the sterilized 0.5cm diameter filter paper on the medium On the surface, drop 10 μl of 2 mg / ml antimicrobial peptide HC14 sample solution dissolved in sterilized deionized water, incubate upside down at 37°C for 18-20 hours, and observe whether the inhibition zone is formed or not. If the sample has antibacterial activity, a clear and transparent bacteriostatic zone will be formed around the filter paper, and the larger the bacteriostatic zone, the stronger the antibacterial activity of the sample.
[0025] (2) Determination of Antimicrobial Peptide HC14 Minimum Inhibitory Concentration (2-fold dilution method):
[0026] Select the strains with the inhibition zone in the previous experiment to carry out th...
Embodiment 3
[0040] HC3-5 stability test:
[0041] Human serum stability test
[0042] Draw 5ml of healthy human blood with a syringe, dispense it into sterilized centrifuge tubes, let it stand at room temperature for 2 hours, and then put it in a refrigerator at 4°C overnight. Centrifuge at 2000rpm for 10min, the light yellow supernatant is human serum. Aspirate the serum with a pipette, aliquot into sterilized centrifuge tubes, and store in a -20°C refrigerator.
[0043] The Hc3-5 antimicrobial peptide sample is mixed with human serum in a certain proportion, so that the concentration of the Hc3-5 antimicrobial peptide sample is 200 μg / ml, the serum concentration is 25%, and the negative control is the mixture of sample solution water and serum. The mixture was incubated in an incubator at 37° C. From 0 to 6 hours, the antibacterial activity of the Hc3-5 antimicrobial peptide against Escherichia coli 1 after serum treatment was detected according to the minimum inhibitory concentration...
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