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738 a.a.
_ZN
 ATP |
+ |
 L-alanine |
+ |
 tRNA(Ala) |
= |
 AMP |
+ |
 diphosphate |
+ |
 L-alanyl-tRNA(Ala) |
|---|
Key reference
DOI no: 10.1073/pnas.0904645106 Proc Natl Acad Sci U S A 106:11028-11033 (2009) PubMed id: 19549823
The structure of alanyl-tRNA synthetase with editing domain. M.Sokabe, T.Ose, A.Nakamura, K.Tokunaga, O.Nureki, M.Yao, I.Tanaka. ABSTRACT
Alanyl-tRNA synthetase (AlaRS) catalyzes synthesis of Ala-tRNA(Ala) and hydrolysis of mis-acylated Ser- and Gly-tRNA(Ala) at 2 different catalytic sites. Here, we describe the monomer structures of C-terminal truncated archaeal AlaRS, with both activation and editing domains in the apo form, in complex with an Ala-AMP analog, and in a high-resolution lysine-methylated form. The structures show docking of the editing domain to the activation domain opposite from the predicted tRNA-binding surface. Thus, the editing site is positioned >35 A from the activation site, prompting us to model 2 different tRNA complexes: one binding tRNA at the activation site, and the other binding tRNA at the editing site. Interestingly, a gel-shift assay also implies the presence of 2 types of tRNA complex with different mobility. These results suggest that tRNA translocation via a canonical CCA flipping is unlikely to occur in AlaRS. The structure also demonstrated the binding of zinc in the editing site, in which the specific coordination of zinc would be facilitated by a conserved GGQ motif, implying that the editing mechanism may not be the same as in ThrRS. As Asn-194 in eubacterial AlaRS important for Ser misactivation is replaced by Thr-213 in archaeal AlaRS, a different Ser accommodation mechanism is proposed.
Selected figure(s)
Figure 2.
Comparison with N453. (A) Superposition of AD of N752 (blue) and N453 (purple), where NX and the regions in N453 corresponding to the deletions in N752 are colored green and orange, respectively. (B) Comparison of HC of N752 (yellow) and N453 (purple) by superposition of HN. α16′ in N453 is colored as orange.Figure 3.
The catalytic sites. (A) The activation site of N752-AlaSA. AlaSA and interacting residues are shown as gray and yellow stick models, respectively. Thr-213 and an interacting water molecule in N752m are superposed as green stick and red ball models (marked as “W”), respectively. Hydrogen bonds in N752-AlaSA and N752m are indicated as black and red dashed lines, respectively. The omit map of AlaSA (at 3.1 Å resolution, contoured at 3 σ) is also shown. (B) Comparison of the editing sites of N752-Zn (yellow) and ThrRS-SerA76 complex (light blue). Only the zinc-binding motifs and SerA76 are shown for ThrRS. The water molecule coordinating to zinc in N752-Zn, and the nucleophile in ThrRS-SerA76 are marked as “W” and “N”, respectively. The GGQ loop is colored green. Interactions in N752-Zn and ThrRS-SerA76 are shown as black and blue dashed lines, respectively.Figures were selected by an automated process.
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