Structural basis of haem-iron acquisition by fungal pathogens

Area of Science:

• Biochemistry
• Structural Biology
• Mycology

Background:

• Pathogenic fungi require iron for survival within host tissues, often facing iron-limited conditions.
• Fungal pathogens utilize secreted haemophores, including those from the Common in Fungal Extracellular Membrane (CFEM) protein family, to scavenge heme-iron from host hemoglobin.
• Understanding the structure and function of these haemophores is critical for developing antifungal strategies.

Purpose of the Study:

• To determine the three-dimensional structure of Csa2, a CFEM protein and haemophore secreted by Candida albicans.
• To elucidate the mechanism by which Csa2 binds and acquires heme, a vital iron source for fungal pathogens.
• To investigate the role of specific amino acid residues, particularly aspartic acid, in heme binding and acquisition.

Main Methods:

• X-ray crystallography was employed to determine the high-resolution 3D structure of the Csa2 haemophore.
• Site-directed mutagenesis was used to create histidine substitution mutants of a key aspartic residue.
• In vitro heme binding assays and in vivo functional studies were conducted to assess mutant protein activity.

Main Results:

• The first 3D structure of a CFEM protein (Csa2) revealed a novel helical-basket fold stabilized by eight cysteines and four disulfide bonds.
• Csa2 binds heme via a unique mechanism involving a hydrophobic platform, an N-terminal handle, and an aspartic residue coordinating Fe3+ specifically.
• Mutants with histidine substitutions at the critical aspartic residue could bind Fe2+ heme and extract heme from hemoglobin but were non-functional in vivo, indicating the importance of oxidation-state-specific binding.

Conclusions:

• The novel structure of the Csa2 haemophore provides insights into heme acquisition by fungal pathogens.
• The aspartic residue's specific coordination of Fe3+ heme is crucial for the functional acquisition of heme in vivo, highlighting a potential target for antifungal drug development.