Molecular basis of alanine discrimination in editing site

AlaX is the homologue of the class II alanyl-tRNA synthetase editing domain and has been shown to exhibit autonomous editing activity against mischarged tRNA^Ala. We solved the structures of AlaX from the archaeon Pyrococcus horikoshii in apo form, complexed with zinc, and with noncognate amino acid L-serine and zinc (Fig. 1). Together with mutational analysis, we demonstrated that the conserved Thr-30 hydroxyl group located near the b-methylene of the bound serine is responsible for the discrimination of noncognate serine from cognate alanine, based on their chemical natures (Fig. 2). Furthermore, we confirmed that the conserved Gln-584 in alanyl-tRNA synthetase, which corresponds to Thr-30 of AlaX, is also critical for discrimination. These observations strongly suggested conservation of the chemical discrimination among trans-and cis-editing of tRNA^Ala.

Fig. 1 The ribbon diagram of the phoAlaX serine–Zn²⁺ complex.

Fig. 2. The editing site. (A) The editing site of PhoAlaX–serine–Zn²⁺ with omit map of serine, zinc, and a water molecule. (B) Schematic representation of the editing site.The red dashed lines indicate the hydrogen bonds with serine. The gray dashed lines indicate the internal hydrogen bonds or the metal coordination that also exist in the structures without serine. The magenta dashed line with the X mark indicates the chemical repulsion. Note that green dashed lines do not represent hydrogen bonds but indicate only distances with each residue. The conserved residues are underlined.

Reference: M. Sokabe, A. Okada, M. Yao, T. Nakashima, I. Tanaka. PNAS, 102, 11669-11674 (2005)