The Mycobacterium tuberculosis (Mtb) heme degradation enzyme, MhuD, has been well characterized both biochemically and structurally. Yet, the fate of MhuD’s tetrapyrrole product, mycobilin, within the cytosol remains unknown. Recent efforts in the Goulding lab have demonstrated that Rv2074, a split β-barrel flavin/deazaflavin oxidoreductases (FDORs), has the ability to reduce mycobilin to a derivative of bilirubin, via a coenzyme F420 dependent mechanism. To further elucidate the mechanism of action of Rv2074, we utilized in silico structural modeling of the Rv2074-mycobilin-F420 complex, which identified potential catalytic residues that play a role in mycobilin binding and/or the reductase activity. Utilizing Rv2074 variants, through kinetic assays and determination of the affinity of mycobilin to each variant, we have shown that certain residues play a role in binding mycobilin, while others play a role in the reductase activity of Rv2074. We are excited to share these results, and to show mechanistic insights into the reductase activity of Rv2074.