31
Molar concentrations of free and bound small molecule
species may be determined from any of a variety of
experimental techniques. The overall stoichiometry of
the process may be obtained from a Scatchard treatment or
a Job's plot. Knowing the overall stoichiometry, initial
values of m and q may be selected which satisfy the simple
relationship mq = n, where n is the total number of DNA
phosphates per bound small molecule. A rapid determination
of the most probable combinations of values may be made by
plotting log ([Pt/m]- q[BHP]) vs.log ( [BHP] / [BH] ) That
pair which yields the most consistent straight line having
a slope of q may then be used to compute values of K for a
series of ratios of [BHP] to [BH]. Again, the pair which
provides the most consistent calculated equilibrium associa
tion constants for all data points, corresponding to either
the surface or intercalative mode, are taken as the exponents
in the binding expression. For interactions between 3-
aminoacridinium cation and calf thymus DNA, best-fit values
of m and q are 3 and 1,respectively for surface binding and
2 and 2 for the intercalative mode (69) To summarize, the
method of Capomacchia allows for the determination of
equilibrium association constants based on mechanistic
considerations. Furthermore, by computing values of K from
data in the midregion of a titration, undue reliance on the
least trustworthy data at the extrema of a titration is
avoided.
Many workers have been satisfied with reporting associ
ation constants calculated from molar quantities rather than