A possible mechanism of processive nucleotide and repeat additions by the telomerase

Abstract

Telomerase is a specialized cellular ribonucleoprotein complex that can synthesize long stretches of a DNA primer by using an intrinsic RNA template sequence. This requires that the telomerase must be able to carry out both nucleotide and repeat additions. Here, based on available structures and experimental data, a model is presented to describe these two addition activities. In the model, the forward movement of the polymerase active site along the template during the processive nucleotide addition is rectified through the incorporation of a matched base, via the Brownian ratchet mechanism. The unpairing of the DNA:RNA hybrid and then repositioning of product 3′-end after each round of repeat synthesis, which are prerequisites for the processive repeat addition, are caused by a force acting on the primer. The force results from the conformational transition of the stem III pseudoknot, which is mechanically induced by the rotation of TERT fingers together with stem IV loop towards the polymerase active site upon a nucleotide binding. Based on the model, the dynamics of processive nucleotide and repeat additions by recombinant Tetrahymena telomerase is studied analytically, which gives good quantitative explanations to the previous experimental results. Moreover, some predicted results are presented. In particular, it is shown that the repeat addition processivity is mainly determined by the difference between the free-energy change required to disrupt the DNA:RNA hybrid and that required to unfold the stem III pseudoknot. A large difference in free energy corresponds to a low repeat addition processivity while a small difference in free energy corresponds to a high repeat addition processivity.