Kinetic studies on ribosomal peptidyltransferase. The behaviour of the inhibitor blasticidin S

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Abstract
In a cell-free system derived from Escherichia coli, the reaction between Ac[3H]Phe-tRNA and puromycin (S) is inhibited by blasticidin S (I). In this reaction Ac[3H]Phe-tRNA is part of the Ac[3H]Phe-tRNA—poly(U)—ribosome complex (C). After preincubating the complex C with I and then adding S, the degree of inhibition is greater than that observed when C reacts with a mixture of S and I. Without preincubation, the inhibition is competitive giving a Ki of 2 × 10−7 M. After preincubation the inhibition becomes of the mixed non-competitive type. A first-order kinetic analysis of the reaction between C and excess S, in the presence or in the absence of I, with or without preincubation, suggests that I acts as a modifier decreasing the catalytic rate constant of ribosomal peptidyltransferase (the putative enzyme that catalyzes the reaction between C and S). The effectiveness of I cannot be expressed by an equilibrium constant such as the above-mentioned Ki. A model is proposed which explains the results obtained. In this model, in the presence of I, C is converted to a modified species C*, which is still able to react with S but with a lower catalytic rate constant. This is a novel concept, in which the ribosome can be subjected to modulation of its activity by small ligands. It can be useful in studies on translational control of protein synthesis.