کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1174412 | 961748 | 2012 | 6 صفحه PDF | دانلود رایگان |
Polyamides (PAs) are distamycin-type ligands of DNA that bind the minor groove and are capable of sequence selective recognition. This capability provides a viable route to their development as therapeutics. Presented here is a simple and convenient fluorescence assay for PA–DNA binding. PAs are titrated into a sample of a hairpin DNA featuring a TAMRA dye attached to an internal dU near the PA binding site. In a study of 6 PAs, PA binding leads to a steady reproducible decrease in fluorescence intensity that can be used to generate binding isotherms. The assay works equally well with both short (6- to 8-ring) and long (14-ring) PAs, and Kd values ranging from approximately 1 nM to at least 140 nM were readily obtained using a simple monochromator or filter configuration. Competition assays provide a means to assessing possible dye interference, which can be negligible. The assay can also be used to determine PA extinction coefficients and to measure binding kinetics; thus, it is an accessible and versatile tool for the study of PA properties and PA–DNA interactions.
Journal: Analytical Biochemistry - Volume 423, Issue 1, 1 April 2012, Pages 178–183