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Direct titration by fluorescent polarization

Direct titration by fluorescent polarization. A = Af - (Ab-Af)[(D+E+Kd)-{(D+E+Kd) 2 -4DE} 1/2 ]/2D. A , anisotropy; Af , anisotropy of the free DNA; Ab , anisotropy of the fully bound DNA; D , the DNA concentration; E , the enzyme concentration. Kd (FAM-AniI wt DNA) = 17.71 ± 2.33 nM

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Direct titration by fluorescent polarization

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  1. Direct titration by fluorescent polarization A = Af - (Ab-Af)[(D+E+Kd)-{(D+E+Kd)2-4DE}1/2]/2D A, anisotropy; Af, anisotropy of the free DNA; Ab, anisotropy of the fully bound DNA; D, the DNA concentration; E, the enzyme concentration. Kd (FAM-AniI wt DNA) = 17.71 ± 2.33 nM Kd (FAM-SceI wt DNA) = 19.78 ± 3.34 nM

  2. Competitive titration X (Bound/Free)= (A-Af)/(Ab-A) C = {1+Ki/(KdX)}[E-D{X/(1+X)}-KdX] A, anisotropy; Af, anisotropy of the free DNA; Ab, anisotropy of the fully bound DNA; C, the unlabled DNA concentration; D, the labled DNA concentration; E, the enzyme concentration. Ki (Unlabled AniI wt DNA) = 51.56 ± 20.37 nM Ki (Unlabled SceI wt DNA) = 120.8 ± 23.18 nM

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