Metal binding motif in the active site of the HDV ribozyme binds divalent and monovalent ions.

ABSTRACT: The hepatitis delta virus (HDV) ribozyme uses both metal ion and nucleobase catalysis in its cleavage mechanism. A reverse G·U wobble was observed in a recent crystal structure of the precleaved state. This unusual base pair positions a Mg(2+) ion to participate in catalysis. Herein, we used molecular dynamics (MD) and X-ray crystallography to characterize the conformation and metal binding characteristics of this base pair in product and precleaved forms. Beginning with a crystal structure of the product form, we observed formation of the reverse G·U wobble during MD trajectories. We also demonstrated that this base pair is compatible with the diffraction data for the product-bound state. During MD trajectories of the product form, Na(+) ions interacted with the reverse G·U wobble in the RNA active site, and a Mg(2+) ion, introduced in certain trajectories, remained bound at this site. Beginning with a crystal structure of the precleaved form, the reverse G·U wobble with bound Mg(2+) remained intact during MD simulations. When we removed Mg(2+) from the starting precleaved structure, Na(+) ions interacted with the reverse G·U wobble. In support of the computational results, we observed competition between Na(+) and Mg(2+) in the precleaved ribozyme crystallographically. Nonlinear Poisson-Boltzmann calculations revealed a negatively charged patch near the reverse G·U wobble. This anionic pocket likely serves to bind metal ions and to help shift the pK(a) of the catalytic nucleobase, C75. Thus, the reverse G·U wobble motif serves to organize two catalytic elements, a metal ion and catalytic nucleobase, within the active site of the HDV ribozyme.

SUBMITTER: Veeraraghavan N 

PROVIDER: S-EPMC3068245 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature