Haloferax volcanii HVO_0665 (HvThi4) is related to members of the THI4 protein family (IPR002922). (A) Multiple amino acid sequence alignment of THI4 homologs including Hfx. volcanii HVO_0665 (HvThi4), Saccharomyces cerevisiae ScTHI4, Arabidopsis thaliana AtTHI4, Thermotoga maritima Tmari_0788, Methanosarcina acetivorans MA_2851, Methanocaldococcus jannaschii MJ0601 and Thermococcus kodakarensis TK0434. Identical and functionally similar amino acid residues are highlighted in black and grey, respectively, with residues conserved with the ScTHI4 Cys205 active site highlighted in red. α helices and β sheets predicted for HVO_0665 by Phyre2-based homology modeling are indicated above the alignment. (B) Cluster analysis of HvThi4 with members of the THI4 protein family. HVO_0665 (HvThi4) of this study is indicated by a circle (●). M. acetivorans MA_2851 and M. jannaschii MJ0601 described as D-ribose-1,5-bisphosphate isomerases and the associated T. kodakarensis TK0434 demonstrated to lack this activity are indicated by squares (■). S. cerevisiae and A. thaliana THI4 enzymes of thiamine biosynthesis are indicated by triangles (▲). Cluster of archaeal THI4 homologs with a conserved active site cysteine residue analogous to ScTHI4 Cys205 are shaded in blue and include uncharacterized proteins of halophilic archaea, Thaumarchaeota, Aeropyrum, and select methanogens and pyrococci. Three letter genus abbreviations are used as proposed by the Subcommittee on the taxonomy of the family Halobacteriaceae. N- and C-termini were trimmed for protein alignments. UniProtKB accession numbers associated with protein sequences are listed in supplemental information.