The role of nitro groups in the binding of nitroaromatics to protein MOPC 315

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Abstract
Two series of dinitrophenyl haptens, in which Cl replaces one or both nitro groups, were used to investigate, by a combination of high-resolution 1H NMR and fluoresence quenching, the presence of groups in the combining site of [mouse myeloma] protein MOPC315, which form H- bonds to the aromatic-ring substituents of the hapten. The large differences in binding constants on successive replacement of nitro groups were due to specific hapten-substituent-protein interactions by showing that there was little difference in the interaction between these haptens and 3-methylindole (a model for the residue tryptophan-93L with which the hapten stacks in protein MOPC315), proving by 1H NMR that the mode of hapten binding is constant and showing that the differences in Kd were consistent with the relative H-bonding capacities of Cl and the nitro moiety. In this way it was established that each nitro group forms a H-bond. From consideration of the 1H NMR chemical shifts of several dinitrophenyl haptens and their trinitrophenyl analogues, there is no distortion of the o-nitro group on binding to the variable fragment of protein MOPC315.