title: Conformational stability and warfarin-binding properties of human serum albumin studied by recombinant mutants.
abstract: Correctly folded recombinant wild-type human serum albumin and the single-residue mutants K199A, W214A, R218H and H242Q were produced with the use of a yeast expression system. The changes in R218H resulted in a pronounced decrease in intrinsic fluorescence. Thermodynamic parameters for thermal denaturation of the present mutants and of five additional mutants have been determined, showing small increases in stability for two mutants (R218H and H242Q) and a larger decrease in stability for one (W214A). In the last of these, denaturation was a heterogeneous process starting at physiological temperature. The high-affinity binding constant for warfarin at pH 7.4 was determined by fluorescence spectroscopy: there was a significant increase in affinity for binding of warfarin to H242Q and K199A and a smaller decrease in affinity for W214A and R218H. The findings show that Trp-214 is not as essential for the high-affinity binding of warfarin as has previously been thought.
authors: Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Kumamoto University, 5-1 Oe-honmachi, Kumamoto 862-0973, Japan.
authors: H
authors: H
authors: Watanabe
authors: U
authors: U
authors: Kragh-Hansen
authors: S
authors: S
authors: Tanase
authors: K
authors: K
authors: Nakajou
authors: M
authors: M
authors: Mitarai
authors: Y
authors: Y
authors: Iwao
authors: T
authors: T
authors: Maruyama
authors: M
authors: M
authors: Otagiri
mesh_terms: D019943:Amino Acid Substitution; D002942:Circular Dichroism; D004795:Enzyme Stability; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012709:Serum Albumin / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D014859:Warfarin / Q000097:blood*
chemical_list: D011994:Recombinant Proteins; D012709:Serum Albumin; D014859:Warfarin