Many pathogenic bacteria require iron for their survival and virulence; in most cases hemin is the main iron source. Pathogens have developed sophisticated heme uptake mechanisms in order to maintain the homeostasis and remain as infectious agents. Corynebacterium diphtheria can obtain hemin during human infection through series of conserved domains (CR) of DxtR-regulated and heme-transport-associated (hta) proteins: HtaA, HtaB, ChtA, ChtB and ChtC. HtaA includes two conserved regions (CR1 and CR2) while the other proteins include a single CR domain. These proteins orchestrate the heme transport to the HmuT protein, a lipoprotein which delivers heme to the ABC membrane transporter HmuUV. Homology modeling of HtaA-CR2, HtaB and ChtB based on amino acid sequence indicated that these proteins have a novel structure. Two tyrosines and one histidine residue are fully conserved in all CR domains. Mutations of these conserved amino acids to alanine significantly lowered the heme binding in comparison to the wild-type proteins. Reconstitution of HtaA-CR2 after removal of the heme with butanone extraction method gave a different form of the protein. UV-visible absorption spectra and resonance Raman spectra data are consistent with heme ligation with an axial tyrosine including a histidine hydrogen-binding partner in HtaA-CR2, HtaB and ChtB. HtaA-CR2 is highly stable to thermal unfolding; the protein was also stable to chemical unfolding using GdnHCl or GdnSCN (up to 4 M at 25^oC) . For HtaA-CR2, unfolding could be observed at 37 ^oC as a single process at high concentrations of denaturant (6.8 – 7.4 M GdnHCl). In contrast, HtaA-CR2 apoproteins (WT, as well as the Y361A and H412A mutants) unfolded readily with low denaturant concentrations (~ 1.3 M GdnHCl). HtaB shows significantly lower stability; a half-life of 330 min was observed in the presence of 6.6 M GdnHCl at 37 ^oC for HtaA-CR2 and a half-life of 39 min was observed in the presence of 4.0 M GdnHCl at 25 ^oC for HtaB; ChtB was very similar to HtaB. For HtaB and ChtB, the high amino acid sequence similarity and identity, similar biophysical characteristics and gene deletion studies suggest that these proteins may function interchangeably during the heme uptake process.