The Use of Orthologous Sequences to Predict the Impact of Amino Acid Substitutions on Protein Function
Figure 4
Accuracy of discrimination between functional and impaired variants by different methods.
Growth-rate metrics for the 30 variants in Table 1 plotted against scores/classifications from various methods that estimate functional impact. The accuracy of each method was determined by calculating the number of mutations correctly called as functional or impaired divided by the number of mutations unambiguously classified by experimental data. Binning of mutations was determined by using a threshold empirically defined by the functional alleles (dashed vertical line in each panel) to define the functional (left of line) and impaired (right of line) bins. (A) SIFT score; note that the graph plots (1–score) to facilitate comparison with the other methods. All functional variants have a SIFT score >0.09 which, when used as a threshold results in a classification accuracy of 62%. The recommended threshold of 0.05 (solid vertical line) results in a lower classification accuracy (42%). (B) Grantham scale of amino acid dissimilarity between wild-type and substituted amino acid. (C) Ancestral Site Preservation (ASP) measure, using inferred ancestral sequences of human MTHFR. Numbers on the x-axis correspond to increasingly ancient ancestors of human MTHFR as defined by the nodes in Figure 3. (D) Ancestral Site Preservation Extended (ASPext) measure. If a site was preserved in the ancestral lineage for a long period before being substituted by the current-day amino acid, the more ancient ancestor is used to define preservation at this site. The preservation measure for 5 variants is shifted by this criterion (see Table 1).
