A kind of antimicrobial peptide gene of African yellow-clawed scorpion, antimicrobial peptide os_116, its structural homologous antimicrobial peptide, preparation method and application
An African yellow-clawed scorpion and antimicrobial peptide technology, applied in the field of bioengineering, can solve the threat of anti-infection treatment and other problems, and achieve the effects of low normal cell toxicity, reduced drug resistance probability, and strong antibacterial activity
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Embodiment 1
[0031] Example 1 Acquisition of Antimicrobial Peptide Gene of African Yellow Clawed Scorpion
[0032] 1. Extraction of total RNA from the venom gland of the African yellow clawed scorpion (RNAiso Plus, purchased from Takara)
[0033] (1) Take the scorpion glands that have been electrocuted and quickly transfer them to a mortar pre-cooled with liquid nitrogen, grind the tissue with a pestle, and add liquid nitrogen continuously until the scorpion glands are ground into powder (the more thorough the grinding, the The higher the yield and quality of RNA), add an appropriate amount of RNAiso Plus that matches the homogenate volume of the sample into the mortar, and fully homogenize;
[0034] (2) Transfer the homogenate to a centrifuge tube and let stand at room temperature (15-30°C) for 5 min;
[0035] (3) Then add chloroform (the volume of chloroform is 1 / 5 of the volume of RNAiso Plus), close the cap of the centrifuge tube tightly, mix until the solution emulsifies and turns mi...
Embodiment 2
[0089] Example 2 Synthesis of the antimicrobial peptide Os_116 from the venom of the African yellow-clawed scorpion
[0090] The amino acid sequence (SEQ ID NO: 2) of the antimicrobial peptide of the African yellow clawed scorpion was deduced according to the antibacterial peptide gene sequence of the African yellow clawed scorpion obtained in Example 1, and the signal peptide and precursor peptide were removed, and the solid phase chemical synthesis method was used. The mature peptide sequence of the African yellow clawed scorpion antimicrobial peptide was synthesized by an automatic peptide synthesizer, and then the synthetic peptide was obtained by desalting and purifying by HPLC reversed-phase C18 column chromatography. The purity of the synthetic polypeptide was identified by high-performance liquid chromatography (HPLC), and its precise molecular weight was determined by matrix-assisted laser desorption ionization time-of-flight mass spectrometry, and the amino acid seque...
Embodiment 3
[0091] Example 3 Design and artificial synthesis of the structurally homologous antimicrobial peptide Os_116 from the venom of the African yellow clawed scorpion
[0092] Structural homologous antimicrobial peptide design of the antimicrobial peptide Os_116 of the venom of the African yellow clawed scorpion: According to the amino acid sequence (SEQ ID NO: 3) and its structural characteristics of the antibacterial peptide Os_116 of the African yellow clawed scorpion venom, the mutants were designed according to the following principles: 1. On the premise of maintaining the α-helical structure of the antimicrobial peptide Os_116 of the venom of the scorpion venom, try to make the amphiphilicity of the mutated antimicrobial peptide more obvious, that is, make its hydrophobic surface more hydrophobic and its hydrophilic surface more hydrophilic ; 2. Enhance the amount of electrostatic charge on the hydrophilic surface and improve its electrostatic attraction to phospholipid membra...
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