The binding of adenosine-14C to polyuridylic acid (poly (U)) and several modified poly-(U)s has been studied by equilibrium dialysis. The poly(U) was modified by addition of appropriate reagents across the 5,6-double bond of the uracil ring to form the photohydrate, photodimer, dihydrouracil, the HOBr addition product and the HSOa-addition product. Modification of the uracil rings decreases the amount of adenosine which can be bound to the poly(U); the decrease in binding is a function of the fraction of uracil rings which have been changed. Using the expression S = So(1 -CYT)~ to relate the fraction of uracil rings modified (T) to the number of binding "sites" remaining (S), it is found that LY is about 1 for all the modifications except photodimer where it is about 2. These observations are taken to mean that the loss of binding capacity of the poly(U) resulting from modifications of the uracil ring is caused by loss of planarity of the uracil rings caused by the modifications, and consequent loss of double helix structure, but that for all modifications except photodimer there is no disruption of the poly(U) double helix on either side of the lesion. There does appear to be local melting on either side of the photodimer lesion. The sigmoidal binding isotherms (Ab versus C,) of modified and unmodified poly(U) can be approximated closely by the following equat.ion:
where Ab = bound A, C. = free A, n = minimum number of adjacent A's in complex, S = concentration of sites on poly(U), and K1 = (Km)ltm for all m 2 n.
The stacking energy of adenosine (w) can be calculated accurately using the following equation, where de/d In C, is obtained from Eq. (1).
(2 )
For unmodified poly(U), w is -2.0 kcal/mole and AGO ( -R T In K1) is -3.2 kcal/mole. The difference (-1.2 kcal/mole) is attributed to hydrogen bonding. Heavily photohydrated poly(U) does not bind guanosine or guanosine-5'-phosphate.