Figure legends
Figure 1 . DsrAB phylogenetic reconstruction showing the taxonomic affiliation of Dsr-hosting organisms and their environmen-tal origin. The Maximum Likelihood reconstruction was conducted on a concatenated alignment of DsrA and DsrB subunits from a curated database comprising the previously recognized primary homolog groups. The scale bar shows the expected substitutions per site. The environmental origin of Dsr homologs is shown based on available metadata associated with the previously published data-base (Muller et al. 2015) or from metadata associated with MAGs from metagenomes. Taxonomic classification is given based on information in the aforementioned database or by >80% amino acid homology to Dsr from cultivars/genomes with taxonomic annotation.
Figure 2. Available Crystal Structures of Dsr and a Homology Model of an Early Evolving Dsr From a Sulfite Reducing Euryarchaeote (MV2). A-C) side view; D-F) top view; A and D) Archaeoglobus. fulgidus heterotetramer – pdb id: 3mmc, B and E) Desulfovibrio. vulgaris hetero-hexamer - pdb id: 2v4j, C and F) homology model of MV2-Eury hetero-hexamer. Chain colors are: A1 (green), B1 (cyan), A2 (light pink), B2 (light blue), C1 and C2 where present (purple and white, respectively). Sirohemes (red), siro-hydrochlorine (salmon), and Fe-S centers (yellow) are sticks or spheres. The sulfite substrate is shown in the D. vulgaris structure (B) in dark blue spheres.
Figure 3 . Close-up view of the “lower” prosthetic group in the structure of Dsr. A) A. fulgidus and B) D. vulgaris Chain colors are : A1 (green), B1 (cyan), Sirohemes (red), siro-hydrochlorine (salmon) and Fe-S centers (yellow) are sticks or spheres. Trp 119 and Thr 135 are shown in spheres and colored by element.
Figure 4 . AlphaFold2 model of MV2.Eury DsrAB. The sequence is colored for pLDDT score (descried in the text), with dark blue corresponding to high confidence prediction (96%) and red to low confidence prediction (43%).The view is a top view of the structure as in Figure 1F. The light blue to red region on the center left and right correspond to residues 50-62 if the A subunit which interact with the DsrC electron donating subunit (Figure 1E, F). The upper left, bottom right and center red regions are the chain termini.
Figure 5 . The heme road FPEC residues in the structure of DsrAB from A. fulgidus . A) Side view B) top view, C) slab of a zoom from the top view Distances between FPEC residues are shown in yellow. Chain colors are: A1 (green), B1 (cyan), A2 (light pink), B2 (light blue). Sirohemes (red sticks) and Fe-S centers (yellow) are also shown. Heme road FPEC residues (blue spheres) tracing a connection between functional hemes are from left to right N180B1, N393A2, T351B1, T351B2, N393A1 and N180B2. N222A1/2 (also blue spheres) interacts with the propionate group of the structural (non-functional) heme visible in (A) in a lower plane. D) Network of aromatic residues connect-ing the residues of the heme road (dark blue) for the left half of the heme road. Three aromatic residues, F317A1, F394A2 and Y348B1, directly connect the heme road residues. These interactions are stabilized from above by N246B1. N246B1 also contacts C244B1 that makes contact with the second Fe-S center distal to the siroheme. Two heme road residues T351B2 and N393A1 of the second heterodimer also appear on the right of the figure.
Figure 6 . Comparison of the heme road FPEC residues in DsrAB. A). D. vulgaris B), the homology model of MV2-Eury. Slab of a zoom from the top views. Chain colors are: A1 (green), B1 (cyan), A2 (light pink), B2 (light blue). Sirohemes (red sticks) and Fe-S centers (yellow) are also shown. FPEC residues (blue spheres) tracing a connection between functional hemes in the D. vulgaris structure (A) are from left to right N191B1 (blue spheres), N410A2 (blue spheres), P368B1 (CPK spheres), P368B2 (CPK spheres), N410A1 (blue spheres) and N191B2 (blue spheres) make up the heme road. The corresponding residues in the MV2.Eury model (B) are from left to right S167B1 (blue spheres), N369A2 (blue spheres), R337B1 (CPK spheres), R337B2 (CPK spheres), N369A1 (blue spheres) and S167B2 (blue spheres). Note that the sulfite ion is shown in darer blue bound to the heme in the D. vulgaris structure (A).
Figure 7 . Internal water molecules in the A1B1 heterodimer of the A. fulgidus structure of DsrAB (A) Full heterodimer and (B) Zoom in the siroheme. Chain colors are: A1 (green), B1 (cyan). Sirohemes (red sticks) and Fe-S centers (yellow) are also shown. Internal water molecules are shown as red spheres. The tryptophan residue blocking access to the structural (lower) heme is shown in CPK spheres.
Figure 8 . Zoom view of the overlaid structures of DsrAB fromA. fulgidus (cyan), D. vulgaris (dark cyan, and MV2 Eury (pale cyan. The DsrB subunit insert in the A. fulgidus structure is in light pink, while the bacterial B loop is magenta. The DsrA subunit insert in the D. vulgaris structure is wheat. The rest of the DsrA1 subunits are green. Some of the FPEC residues (K and F/D/F) are indicated in red stick and distances between them (7.4 and 26.7 Å are indicated). The lysine residues of the FPEC form a salt bridge with a glutamate from the DsrA subunit (2.7 Å).
Figure 9 . DsrAB phylogenetic reconstruction showing the taxonomic affiliation of Dsr-hosting organisms and the presence of DsrA and DsrB structural inserts. The Maximum Likelihood reconstruction was conducted on a concatenated alignment of DsrA and DsrB subunits from a curated database comprising the previously recognized primary homolog groups. Scale bar shows the expected substitutions per site. Taxonomic classification is given based on information in the database (Muller et al. 2015) or by >80% amino acid homology to Dsr from cultivars/genomes with taxonomic annotation. DsrA or DsrB inserts were identified based on structural characterizations and subsequent identification within DsrA or DsrB alignments.