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Paradox of mistranslation of serine for alanine caused by AlaRS recognition dilemma


Mistranslation arising from confusion of serine for alanine by alanyl-tRNA synthetases (AlaRSs) has profound functional consequences1,2,3. Throughout evolution, two editing checkpoints prevent disease-causing mistranslation from confusing glycine or serine for alanine at the active site of AlaRS. In both bacteria and mice, Ser poses a bigger challenge than Gly1,2. One checkpoint is the AlaRS editing centre, and the other is from widely distributed AlaXps—free-standing, genome-encoded editing proteins that clear Ser-tRNAAla. The paradox of misincorporating both a smaller (glycine) and a larger (serine) amino acid suggests a deep conflict for nature-designed AlaRS. Here we show the chemical basis for this conflict. Nine crystal structures, together with kinetic and mutational analysis, provided snapshots of adenylate formation for each amino acid. An inherent dilemma is posed by constraints of a structural design that pins down the α-amino group of the bound amino acid by using an acidic residue. This design, dating back more than 3 billion years, creates a serendipitous interaction with the serine OH that is difficult to avoid. Apparently because no better architecture for the recognition of alanine could be found, the serine misactivation problem was solved through free-standing AlaXps, which appeared contemporaneously with early AlaRSs. The results reveal unconventional problems and solutions arising from the historical design of the protein synthesis machinery.

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Figure 1: Plasticity of active site in AlaRS.
Figure 2: Alanine active site with intrinsic design defect that mis-binds serine.
Figure 3: Mutant enzyme with a smaller pocket retains serine binding affinity.
Figure 4: Critical role of Asp 235 shows dilemma for AlaRSs.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors for the reported crystal structures have been deposited with the Protein Data Bank under accession codes 3hxu (WT/Ala-SA), 3hxv (WT/Gly-SA), 3hxw (WT/Ser-SA), 3hxx (WT/AMP-PCP/Mg(II)), 3hxy (WT/Ala-AMP/PCP/AMP-PCP/Mg(II)), 3hxz (G237A/Ala-SA), 3hy0 (G237A/Gly-SA) and 3hy1 (G237A-apo and G237A/Ser-SA).


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We thank R. J. Read for pointing out the errors in the previous AlaRS–ligand structures and for crystallographic discussions; and G. J. Kleywegt, Z. Otwinowski and A. Perrakis for technical assistance. X-ray diffraction data were collected at Stanford Synchrotron Radiation Laboratory (SSRL) beamlines 7-1, 9-1 and 11-1. This work was supported by grant GM 15539 from the National Institutes of Health and by a fellowship from the National Foundation for Cancer Research.

Author Contributions M.G., X.-L.Y. and P.S. designed the experiments. M.G., Y.E.C., R.S. and K.B. performed the experiments. M.G. and Y.E.C. analysed the data. M.G., Y.E.C., X.-L.Y. and P.S. wrote the paper. All authors discussed the results and commented on the manuscript.

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Correspondence to Paul Schimmel.

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Supplementary Figures

This file contains Supplementary Figures 1-8 with Legends. (PDF 5485 kb)

Supplementary Table 1

This file contains a summary of the data collection and refinement statistics of the nine structures. (PDF 192 kb)

Supplementary Movie 1

This movie file shows that serine paradox is caused by AlaRS recognition dilemma (see Supplementary Figures file for full Legend). (MOV 6540 kb)

Supplementary Movie 2

This movie file shows the alanyl-adenylate formation mechanism (see Supplementary Figures file for full Legend). (MOV 4436 kb)

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Guo, M., Chong, Y., Shapiro, R. et al. Paradox of mistranslation of serine for alanine caused by AlaRS recognition dilemma. Nature 462, 808–812 (2009).

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