H. influenzae is a Gram-negative, human restricted commensal of the upper respiratory tract, and a significant opportunistic pathogen of children under five. Type b H. influenzae infection can result in meningitis, epiglottitis, septic arthritis, cellulitis, and pneumonia, and can be fatal. Moreover, nontypeable H. influenzae (NTHi), which is not covered by the current H. influenzae type b polysaccharide-protein conjugate vaccine, can cause acute otitis media, sinusitis, conjunctivitis, chronic obstructive pulmonary disease, septiceaemia and pneumonia, and is increasingly becoming more prevalent in countries with high H. influenzae type b vaccination rates. An increased prevalence of multidrug resistant (MDR) and extensively drug resistant (XDR) H. influenzae isolates have also been reported in recent years, necessitating further investigation into potential therapeutic targets that could cover all H. influenzae types and levels of resistance. In order to survive in the host, H. influenzae must scavenge iron from host iron-containing proteins either via siderophores or TonB-dependent transporters present in the outer membrane. As such, we sought to further characterize the structural and biochemical basis of heme acquisition of an understudied haemoglobin-haptoglobin TonB transporter, Hgp2. Due to poor results from expression trials of Hgp2 in E. coli, we optimised production and purification of Hgp2 in an avirulent H. influenzae strain, using a Pasteurella vector with Strep-Tag cassette under control of a native promoter. From this, we were able to probe the specific interactions between Hgp2 and haemoglobin-haptoglobin using multiple biochemical methods, including differential scanning fluorimetry, surface plasmon resonance, and cryo electron microscopy. Our results reveal crucial insight into the molecular mechanisms underpinning Hgp2-haemoglobin-haptoglobin binding.