Loontiens F G, McLaughlin L W, Diekmann S, Clegg R M
Abteilung Molekulare Biologie, Max Planck Institut für biophysikalische Chemie, Göttingen, FRG.
Biochemistry. 1991 Jan 8;30(1):182-9. doi: 10.1021/bi00215a027.
Fluorescence titrations have been carried out to determine the association constants (Ka) for binding of the dyes Hoechst 33258 and DAPI to the self-complementary decamer d(CTGAATTCAG) and nine duplex derivatives with exocyclic substituent changes in the six central base pairs. Many Ka values are in the range (2-5) x 10(8) (duplex M)-1 at 5.5 degrees C. Replacement of the leftmost adenine by 2-aminopurine in the sequence decreases Ka for Hoechst 33258 by a factor of 170. When the centermost adenine is replaced by 2-aminopurine, Ka for Hoechst 33258 and DAPI is too small to be evaluated. When the centermost adenine is replaced by purine, Ka for both dyes increases, but this very stable duplex-Hoechst 33258 complex is nonfluorescent. The measured affinities are compared to expectations derived from X-ray studies with dodecamer-dye complexes having an identical central binding sequence (Pjura et al., 1987; Teng et al., 1988; Larsen et al., 1989).
已进行荧光滴定以确定染料Hoechst 33258和DAPI与自身互补的十聚体d(CTGAATTCAG)以及六种中心碱基对带有环外取代基变化的九种双链体衍生物结合的缔合常数(Ka)。在5.5℃时，许多Ka值在(2 - 5)×10⁸(双链体M)⁻¹范围内。序列中最左边的腺嘌呤被2-氨基嘌呤取代会使Hoechst 33258的Ka降低170倍。当中心腺嘌呤被2-氨基嘌呤取代时，Hoechst 33258和DAPI的Ka太小而无法评估。当中心腺嘌呤被嘌呤取代时，两种染料的Ka均增加，但这种非常稳定的双链体 - Hoechst 33258复合物是无荧光的。将测得的亲和力与来自具有相同中心结合序列的十二聚体 - 染料复合物的X射线研究结果进行比较(Pjura等人，1987年；Teng等人，1988年；Larsen等人，1989年)。
