Cooperative interactions between the amino- and carboxyl-terminal lobes contribute to the unique iron-binding stability of lactoferrin.
Lactoferrin is a member of the transferrin family of iron-binding proteins. Several functions have been ascribed to lactoferrin, including regulation of iron homeostasis, antibacterial properties, and regulation of myelopoiesis. However, the structural features of lactoferrin that are required for most of these functions are unknown. Previously, we reported the development of an efficient fungal expression system to produce recombinant human lactoferrin. The availability of this production system demonstrated the feasibility of producing mutant lactoferrins to address the structure/function relationship of the protein. In the present study, we used a site-directed mutagenesis approach to address the contribution of the bilobal structure of lactoferrin to its unique iron-binding stability. Like transferrin, lactoferrin consists of two repeated iron-binding lobes that bind one iron atom each. However, unlike transferrin, lactoferrin retains iron over a broad pH range, a key property that contributes to the unique iron-binding functions of the protein. Using mutants that selectively ablate the iron-binding function in either lobe, we demonstrate differential iron-binding stability of the amino- and carboxyl-terminal iron-binding lobes of lactoferrin. Further, we show that the unique iron-binding stability of the protein is imparted primarily by the carboxyl-terminal domain which functions cooperatively to stabilize iron-binding to the amino-terminal domain of lactoferrin.
Ward
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Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
Zhou
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Conneely
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D001483:Base Sequence; D002460:Cell Line; D006801:Humans; D007781:Lactoferrin / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D007781:Lactoferrin