Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, requires iron for survival. In Mtb, MhuD is the cytosolic protein that degrades imported heme. MhuD is distinct, in both sequence and structure, from canonical heme oxygenases (HOs) but homologous with IsdG-type proteins. Canonical HO is found mainly in eukaryotes, while IsdG-type proteins are predominantly found in prokaryotes, including pathogens. While there are several published structures of MhuD and other IsdG-type proteins in complex with the heme substrate, no structures of IsdG-type proteins in complex with a product have been reported, unlike the case for HOs. We recently showed that the Mtb variant MhuD-R26S produces biliverdin IXα (αBV) rather than the wild-type mycobilin isomers. Given that mycobilin and other IsdG-type protein products like staphylobilin are difficult to isolate in quantities sufficient for structure determination, here we use the MhuD-R26S variant and its product αBV as a proxy to study the IsdG-type protein-product complex. First, we show that αBV has a nanomolar affinity for MhuD and the R26S variant. Second, we determined the MhuD-R26S-αBV complex structure to 2.5 Å, which reveals two notable features: (1) two αBV molecules bound per active site and (2) a novel α-helix (α3) that was not observed in previous MhuD-heme structures. Finally, through molecular dynamics simulations, we show that α3 is stable with the proximal αBV alone. MhuD's high affinity for the product and the observed structural and electrostatic changes that accompany substrate turnover suggest that there may be an unidentified class of proteins that are responsible for the extraction of products from MhuD and other IsdG-type proteins.

Upregulation of interleukin-17 receptor B (IL-17RB) is known to be oncogenic, while other IL-17 receptors and ligands are generally involved in pro-inflammatory pathways. We identify a mouse neutralizing monoclonal antibody (mAb) D9, which blocks the IL-17RB/IL-17B pathway and inhibits pancreatic tumorigenesis in an orthotopic mouse model. The X-ray crystal structure of the IL-17RB ectodomain in complex with its neutralizing antibody D9 shows that D9 binds to a predicted ligand binding interface and engages with the A'-A loop of IL-17RB fibronectin III domain 1 in a unique conformational state. This structure also provides important paratope information to guide the design of antibody humanization and affinity maturation of D9, resulting in a humanized 1B12 antibody with marginal affinity loss and effective neutralization of IL-17B/IL-17RB signaling to impede tumorigenesis in a mouse xenograft model.