Fig. 5

Structural mapping of disease-related variants in WARS1 and SARS1 and functional characterization of the WARS1 variant. (a) Upper panel: structure of the human WARS1 homodimer (PDB 2DR2). Protein and RNA are shown as cartoon. One monomer is colored according to domains as depicted in Figure 2d. The other monomer is shown in grey. The tRNA and Arg133 are shown in red. Arg133 is shown as sphere in the overview. The region containing Arg133 is shown enlarged, with important residues shown as sticks. Lower panel: structure of human SARS1 in complex with Ser-SA (PDB 4L87). The protein is shown as cartoon and domains colored in accordance with Figure 2d. Ser-SA is shown as sticks and Arg302 and Arg390 are shown as red spheres. The active site is shown enlarged with Arg302 and Arg390 shown as sticks and Mg ions and phosphate shown as spheres. (b) Densitometric quantification of WARS1 protein abundance, relative to β-actin loading control, in patient fibroblasts from individual 2 (I2) and three control fibroblasts (C1, C2, C3) (n = 6). Results presented as boxplots (center line represents median, upper and lower hinge show 25th and 75th percentiles, upper and lower whisker extend to largest/smallest value within 1.5× inter-quartile range of the respective hinge, additionally all datapoints are presented individually). Results of ANOVA and post hoc Tukey analysis are presented as compact letter display of all pair-wise comparisons in increasing order. Lower panel: representative WARS1 immunoblots of three replicates each and loading controls (β-actin). (c) Aminoacylation activity was normal in fibroblasts from individual 2 (I2) compared to two control fibroblasts and endogenous activity of aminoacyl tRNA-transferases YARS1 and FARS1. ANOVA, analysis of variance; mRNA, messenger RNA; tRNA, transfer RNA.