Enzyme Structure with Two Catalytic Sites for Double-Sieve Selection of Substrate

TLDR

High‑fidelity genetic information transfer relies on editing reactions, and a double‑sieve mechanism has been proposed for two‑step substrate selection. The authors present a 2.5‑Å crystal structure of an isoleucyl‑tRNA synthetase with editing activity. The enzyme activates both l‑isoleucine and the similar l‑valine during aminoacylation, then rapidly hydrolyzes only valylated products in a second editing step, and the crystal structures reveal the first sieve in the Rossmann‑fold aminoacylation domain and the second sieve in a protruding β‑barrel editing domain. The crystal structures confirm that the first sieve resides in the aminoacylation Rossmann fold and the second sieve in a globular β‑barrel editing domain.

Abstract

High-fidelity transfers of genetic information in the central dogma can be achieved by a reaction called editing. The crystal structure of an enzyme with editing activity in translation is presented here at 2.5 angstroms resolution. The enzyme, isoleucyl–transfer RNA synthetase, activates not only the cognate substrate l -isoleucine but also the minimally distinct l -valine in the first, aminoacylation step. Then, in a second, “editing” step, the synthetase itself rapidly hydrolyzes only the valylated products. For this two-step substrate selection, a “double-sieve” mechanism has already been proposed. The present crystal structures of the synthetase in complexes with l -isoleucine and l -valine demonstrate that the first sieve is on the aminoacylation domain containing the Rossmann fold, whereas the second, editing sieve exists on a globular β-barrel domain that protrudes from the aminoacylation domain.