title: Mutants and molecular dockings reveal that the primary L-thyroxine binding site in human serum albumin is not the one which can cause familial dysalbuminemic hyperthyroxinemia.
abstract: BACKGROUND
  Natural mutations of R218 in human serum albumin (HSA) result in an increased affinity for L-thyroxine and lead to the autosomal dominant condition of familial dysalbuminemic hyperthyroxinemia.
  
  
  METHODS
  Binding was studied by equilibrium dialysis and computer modeling.
  
  
  RESULTS
  Ten of 32 other isoforms tested had modified high-affinity hormone binding. L-thyroxine has been reported to bind to four sites (Tr) in HSA; Tr1 and Tr4 are placed in the N-terminal and C-terminal part of the protein, respectively. Site-directed mutagenesis gave new information about all the sites.
  
  
  CONCLUSIONS
  It is widely assumed that Tr1 is the primary hormone site, and that this site, on a modified form, is responsible for the above syndrome, but the binding experiments with the genetic variants and displacement studies with marker ligands indicated that the primary site is Tr4. This new assignment of the high-affinity site was strongly supported by results of MM-PBSA analyses and by molecular docking performed on relaxed protein structure. However, dockings also revealed that mutating R218 for a smaller amino acid increases the affinity of Tr1 to such an extent that it can become the high-affinity site.
  
  
  CONCLUSIONS
  Placing the high-affinity binding site (Tr4) and the one which can result in familial dysalbuminemic hyperthyroxinemia (Tr1) in two very different parts of HSA is not trivial, because in this way persons with and without the syndrome can have different types of interactions, and thereby complications, when given albumin-bound drugs. The molecular information is also useful when designing drugs based on L-thyroxine analogues.
authors.affiliation: Department of Biomedicine, University of Aarhus, DK-8000 Aarhus C, Denmark. Electronic address: ukh@biomed.au.dk.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Kragh-Hansen
authors.affiliation: Department of Molecular Medicine, University of Pavia, I-27100 Pavia, Italy.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Minchiotti
authors.affiliation: Department of Chemistry and Interdisciplinary Nanoscience Center, University of Aarhus, DK-8000 Aarhus C, Denmark.
authors.forename: Andrea
authors.initials: A
authors.lastname: Coletta
authors.affiliation: Department of Molecular Biology & Genetics and Interdisciplinary Nanoscience Center, University of Aarhus, DK-8000 Aarhus C, Denmark.
authors.forename: Konrad
authors.initials: K
authors.lastname: Bienk
authors.affiliation: Department of Molecular Medicine, University of Pavia, I-27100 Pavia, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Galliano
authors.affiliation: Department of Chemistry and Interdisciplinary Nanoscience Center, University of Aarhus, DK-8000 Aarhus C, Denmark.
authors.forename: Birgit
authors.initials: B
authors.lastname: Schiøtt
authors.affiliation: Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Iwao
authors.affiliation: Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Ishima
authors.affiliation: Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Otagiri
mesh_terms: D001665:Binding Sites; D050010:Hyperthyroxinemia, Familial Dysalbuminemic; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D012709:Serum Albumin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013974:Thyroxine / Q000737:chemistry* / Q000378:metabolism
chemical_list: D012709:Serum Albumin; D013974:Thyroxine
keywords: Computer modeling; Genetic variants; High-affinity binding site; Human serum albumin; L-thyroxine; Site-directed mutagenesis