Gain-of-function experiments with bacteriophage lambda uncover residues under diversifying selection in nature

Maddamsetti, Rohan; Johnson, Daniel T.; Spielman, Stephanie J.; Petrie, Katherine L.; Marks, Debora S.; Meyer, Justin R.

We used the evcouplings pipeline (Hopf et al. 2017) to generate jackhmmer (Eddy 2011) alignments of 1207 full-length J protein homologs (parameter settings: bitscore = 0.2; theta = 0.999; seqid filter = 95 and 99 (thresholds for filtering highly similar sequences); minimum sequence coverage = 99 (require sequences to align to 99% of wild-type J protein); and minimum sequence coverage = 50). By default, this alignment only includes residues found in the wild-type J protein and excludes insertions relative to wild-type J. We included these insertions by querying full-length sequences in a larger Stockholm-formatted alignment produced by the evcouplings pipeline. When analyzing the specificity region (say, constructing a phylogeny), we used residues 960-1132 of this full-length alignment.