APC/Asef多肽抑制剂结构的优化及生物学活性研究

Structure optimum and biological activity studies of APC/Asef peptide inhibitors

目的·设计并合成结肠腺瘤息肉易感基因(adenomatous polyposis coli,APC)/鸟苷酸交换因子(APC-stimulated guanine nucleotide exchange factor,Asef)多肽抑制剂,探讨该抑制剂不同构效之间的关系。方法·基于APC-MAI-150复合物结构,一方面在MAI-150多肽N端用3-苯基丙酰(3-phenylpropane)、Glu-Glu(GG)和β-Ala-Ala(β-AA)取代连氨基基团苄氧羰基(carbobenzoxy,CBZ),另一方面尝试将MAI-150中N端第6位Tyr侧链苯环上的对位羟基替换为-CN、-NO_2、-NH_2和-F,设计合成新的7条多肽。荧光偏振活性检测多肽亲和力,并根据活性结果将多肽MDL-3、MDL-4和MDL-5对接到APC蛋白中,联合计算机对接的多肽和APC蛋白的结合模式,探讨该3条多肽的构效关系。结果·新合成的7条多肽中,多肽MDL-5的N端3-phenylpropane连氨基亲和力最高,其半数有效抑制浓度(half maximal inhibitory concentration,IC50)为2.35μmol/L;与MDL-5相比,多肽N端用GG(MDL-6)和β-AA(MDL-7)连接氨基亲和力明显降低;MDL-3和MDL-4的N端第6位Tyr侧链苯环上的对位羟基替换为-NH_2和-F亲和力相比较于替换为-CN(MDL-1)和-NO_2 (MDL-2)要明显提高;但新合成的多肽亲和力均低于MAI-150。结论·改造多肽N端连氨基基团和第6位Tyr侧链苯环上的对位羟基无助于提高多肽的亲和力。

Objective·To design and synthesize APC/Asef peptide inhibitors,and investigate the relationship between the structures and affinity of peptides.Methods·Based on crystal structure of the APC-MAI-150 complex,on the one hand,3-phenylpropane,Glu-Glu(GG)andβ-Ala-Ala(β-AA)were used to replace carbobenzoxy(CBZ)at the N terminal of MAI-150 to bind-NH2;on the other hand,-CN,-NO2,-NH2,and-F were replaced the parahydroxyl on the sixth tyrosine(Tyr)side chain benzene at the N terminal of MAI-150.And seven peptides were synthesized.Fluorescence polarization was applied to test peptide affinity,and molecular design laboratory-3(MDL-3),MDL-4 and MDL-5 were docked into APC protein based on the results of activity.The structure-activity relationship of the three peptides was studied by combining the binding patterns of computer docked peptide and APC protein.Results·Among the seven peptides,the N terminal 3-phenylpropane linked-NH2 of peptide MDL-5 had the highest affinity,which IC50 was 2.35μmol/L.Compared with MDL-5,the affinity of MDL-6 and MDL-7 were significantly reduced.The para-hydroxyl on the sixth Tyr side chain benzene at the N terminal replaced with-NH2(MDL-3)and-F(MDL-4),which the affinity were higher than-CN(MDL-1)and-NO2(MDL-2).However,the affinity of newly synthesized peptides was lower than MAI-150.Conclusion·Transforming peptide N terminal linked-NH2 and the para-hydroxyl group on the sixth Tyr side chain benzene doesn't help to improve the affinity of peptides.