Fig. 1.

Basis for distinguishing e-pilins from other pilins. Top: Alignment showing the location of aromatic residues in each pilin tested for conductivity in previous studies (Table S1). Dark horizontal lines indicate 42–53 aa aromatic-free gaps in non-conductive pilins. Conserved N-terminal aromatic residues in bacterial e-pilins are indicated by vertical boxes. All bacterial e-pilins contained F-1, Y-24, Y-27, Y/F-51, Y/F-50 and/or Y/F-51, and H/Y/F-32 and/or Y/F-57. The only N-terminal residues shared by archaeal and bacterial e-pilins were F-1 and Y-57. Bottom: Relationship between gap size and percentage of aromatic amino acids in the mature pilin peptide for four types of pilins, which was used to establish conservative criteria for identifying putative e-pilins in environmental metagenomes. Therefore, we used ≥9.8% aromatics and ≤22-aa aromatic-free gap (boxed area labelled ‘conductive e-pilins’), and the presence of aromatic amino acids at residues as a conservative threshold for predicting putative e-pilins from metagenomes, consistent with thresholds established by Walker et al. (2019a). Additional information about each pilin is in Tables S1 and S2.