丙酸睾酮对大肠杆菌RecQ解旋酶结构和功能的影响

Effects of testosterone propionate on the structure and function of E.coli RecQ helicase

目的研究体外丙酸睾酮对大肠杆菌RecQ解旋酶结构和功能的影响及二者的相互作用机制。方法分别运用荧光偏振技术、ATPase试剂盒和紫外吸收光谱技术分析丙酸睾酮对大肠杆菌RecQ解旋酶活性和DNA结合活性、AT-Pase活性和构象的影响。结果丙酸睾酮能够影响E.coliRecQ解旋酶的活性。低浓度促进解旋酶活性,高浓度抑制;抑制RecQ解旋酶与dsDNA的结合活性,其抑制常数Ki小于7×10-5 nmol.L-1;对与ssDNA结合活性的Ki为7×10-3 nmol.L-1;对ATPase活性抑制作用的时间相关性较弱;影响RecQ解旋酶的构象变化,但峰位无偏离。结论提示丙酸睾酮与RecQ解旋酶具有弱相互作用,二者具有两个结合位点。

Aim To study the effects of testosterone propionate on the bioactivity and structure of E.coli RecQ helicase,and the interaction molecular mechanisms in vitro.Methods The effects of the testosterone propionate on the helicase activity,DNA binding activity,ATPase activity,and the structure of E.coli RecQ helicase were assayed by fluorescence polarization,PiColorLock Gold reagent,and ultraviolet aborption spectrum respectively.Results The biological activity and structure of E.coli RecQ helicase were weakly effected by testosterone propionate.The helicase activity was promoted at low concentration and inhibited at high concentration,the Ki for DNA binding activity was below 7×10-5 nmol·L-1 when sustrate was dsDNA,but the inhibition was very weak when sustrate was ssDNA,and the Ki at least 7×10-3 nmol·L-1.RecQ helicase UV absorption peak was changed by testosterone propionate,but the maximum UV absorption peak would not deviate.Conclusions The interaction between E.coli RecQ helicase and testosterone propionate was weak.The binding sites between testosterone propionate and E.coli RecQ helicase had two varieties.