Structural basis of haptoglobin-haemoglobin scavenging by CD163

Thesis

Haemoglobin (Hb) is essential for oxygen transport in vertebrates. However, when it is released from red blood cells, its haem group becomes reactive and must be tightly regulated to prevent its toxicity. Haptoglobin (Hp) acts as the first line of defence by scavenging Hb in the serum. Humans produce three major Hp phenotypes, the dimeric Hp 1-1 as well as the multimeric Hp 2-1 and 2-2. After complexing with Hb, the scavenger receptor CD163 of macrophages mediates the endocytosis of the haptoglobin-haemoglobin complex (HpHb), resulting in its detoxification. Here, I present the structural basis of HpHb scavenging by CD163. Cryogenic electron microscopy reveals that CD163 forms dimers and trimers, with the protomers collectively binding to a single ‘head’ of HpHb in an asymmetric fashion. The receptor primarily interacts with Hb but also contacts Hp. Uptake experiments demonstrate that this allows CD163 to mediate preferential uptake of HpHb, while Hb is internalised at a lower rate. Spheres of atomic density are present at both the ligand binding and multimerisation interfaces, and these are attributed to calcium. Indeed, biophysical assays show that calcium ions are critical for ligand binding. Mutational studies demonstrate that while monomeric CD163 can scavenge the multimeric Hp(2-2)Hb, uptake of the dimeric Hp(1-1)Hb and Hb is impaired compared to the wild-type receptor, suggesting that receptor multimerisation evolved to facilitate uptake of lower affinity ligands. I also present the cryo-electron microscopy structure of unliganded CD163. In the dimeric form, CD163 exhibits an ‘arm-to-arm’ contact, suggesting a potential regulatory mechanism. These results provide novel insights into the molecular mechanisms governing HpHb scavenging, shedding light on how CD163 selects its ligand and highlighting the important roles of calcium and multimerisation.

Authors: Zhou, X

• DOI: 10.5287/ora-ga8emne2j
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: ligand uptake; CD163; haptoglobin; haemoglobin