Extended Data Fig. 1: Orthogonal validation of growth rate inference, structural explanation for antitoxin mutation effects, and covariational signal between toxin–antitoxin ParE3/ParD3.

a, Comparison of growth rates inferred by high-throughput vs. individual growth measurement. X axis error bars indicate + /− 2x standard deviation derived from n = 10 or n = 11 technical plate reader replicates (see Methods). Y axis error bars indicate 95% posterior highest density interval. The Pearson correlation coefficient (r) is indicated. b, Raw log-read ratio reproducibility between replicates (+1 pseudocount) for all single and double mutants. The Pearson correlation coefficient (r) is indicated. c, Mean mutation effect of residues in the C-terminal α-helix 3 of the ParD3 antitoxin indicates that residues facing the toxin are more susceptible to mutations that disrupt the ParD3–ParE3 interaction, producing negative Δgrowth rate values. d, Mean mutation effect in the N-terminal oligomerization region of the antitoxin highlights residues susceptible to disrupting the ParD3–ParE3 interaction when mutated. Cartoon illustrates arrangement of ParE3–ParD3 octamer observed in the co-crystal structure (PDB: 5CEG). One of the 4 antitoxin monomers is coloured by the mean mutation effect. e, Top 10 toxin–antitoxin covarying residue pairs indicated for reference. f, The 90% precision cutoff yields 29 toxin–antitoxin covarying residue pairs (black in upper, right quadrant) of which 28 pairs fall within toxin–antitoxin interface residues that are < 6 Å minimum atom distance (ochre dots) in the ParE3-D3 crystal structure (PDB ID: 5CEG).