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Higher order interaction terms (mutation pairs) were included to account for synergistic and antagonistic effects between mutations.
Mutation pairs were ranked by physical distance.
The estimated deviations obtained using this particular subset of mutation pairs were also most successful in predicting the functional relationships (AUC = 0.810).
The order in which the mutation pairs were included into the approximation process further improved the relative classification power (AUC = 0.789, p = 0.002).
Perhaps the biggest technical limitation of the present work concerns the heuristic way in which the subsets of mutation pairs were selected for the approximation of the double-mutant fitness matrix.
Correlated mutation pairs were compared with protein structure data as follows.
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The p-value is calculated for the null hypothesis that the covariation scores for the two types of mutation pairs are from the same distribution.
The subset of mutation pairs being used in the approximation process had a marked effect on the distribution of the estimated deviations (Figure 4).
Correlated mutation pairs are grouped based on their connectivity to each other; only those "correlated groups" were analyzed.
Overall, 63% of the correlated mutation pairs are conserved in at least one other MADS protein, and 37% are not (conserved intramolecular correlated mutations are listed in Additional File 7).
However, the compensatory mutation pair is not coincided in the study.
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