Structure And Analgesic Activity Study Of BmK AGP-SYPU1 From Buthus Martensii Karsch

BmK AGP-SYPU1 is a kind of analgesic peptide which is purified from the venom of Chinese scorpion Buthus martensii Karsch(BmK),the result suggests BmK AGP-SYPU1 can inhibit guts pain of mouse by using mouse-twisting test.Analyzed the relationship of Structure and analgesic function of BmK AGP-SYPU1,the results predict that analgesic function domain maybe consist of molecular positive domain,negative domain and Face A(hydrophobic domain),analgesic activity maybe has relationship with amino acid Q8,Y5,Y10,Y42,R18,K62 and H64 and so on.Using SOE PCR,fifteen site-directed mutagenesis of BmK AGP-SYPU1 had been built.The recombinant expression vectors are pSYPU-1b-Tag(His)-Mut Y5F,pSYPU-lb-Tag(His)-Mut Y5A,pSYPU-1b-Tag(His)-Mut Y5W,pSYPU-lb-Tag(His)-Mut Q8A,pSYPU-lb-Tag(His)-Mut Q8K,pSYPU-lb-Tag(His)-Mut Y10H,pSYPU-lb-Tag(His)-Mut R18K,pSYPU-lb-Tag(His)-Mut Y42F,pSYPU-lb-Tag(His)-Mut Y42S,pSYPU-1b-Tag(His)-Mut Y42T,pSYPU-lb-Tag(His)-Mut Y42G,pSYPU-lb-Tag(His)-Mut Y5WY42F,pSYPU-lb-Tag(His)-Mut K62L,pSYPU-lb-Tag(His)-Mut H64K and pSYPU-1b-Tag(His)-Mut H64R.The recombinant expression vectors were transformed into E.coli BL21(DE3)and efficiently expressed as a soluble and functional peptide.They were purified by metal chelating chromatography and cation exchange chromatography.About 1～2mg/L purified recombinant BmK AGP-SYPU1 could be obtained.Analgesic activity assays of mice suggest that the amino acid Y5,Y10,R18 and Y42 of BmK AGP-SYPU1 obviously affect analgesic activity.The analgesic function domain consists of molecular a positive domain,a negative domain and hydrophobic domains.Y5 and Y42 obviously affect analgesic activity.The interaction between amino acid residues Y5 and Y42 and between Y5 and the main chain of β-turm3 affects the spatial trends of β-turn3.The location of P-turn 3 affects analgesic activity of the peptide.When β-turn 3 turn to middle of molecular,analgesic activity of the peptide has been increased,otherwise,analgesic activity of the peptide has been decreased.