Ligand-induced conformation change in folate-binding protein.
C.d. and fluorescence spectroscopy have been used to investigate the effect of ligand binding on the structure and stability of folate-binding protein (FBP) from cow's whey. The c.d. spectrum of unligated FBP predicts the following secondary structure: 22% helix, 25% antiparallel beta-strand, 5% parallel beta-strand, 17% turn and 31% random-coil structure. Folate binding to FBP results in significant changes in the c.d. spectrum. Analysis of the spectrum shows a 10% decrease in antiparallel beta-strand as a result of ligand binding. Folate binding also leads to strong quenching of FBP tryptophan fluorescence. The magnitude of the quench is proportional to ligand binding. The guanidinium chloride-induced unfolding of FBP is shown to be a multistate process. Detection by c.d. and fluorescence spectroscopy lead to non-identical transitions. Modelling studies are consistent with the existence of a stable folding intermediate. Ligand binding to FBP increases the apparent folding stability of the molecule. Simultaneous detection by c.d. and fluorescence indicate that the apparent increased folding stability is derived from ligand-induced aggregation of FBP.
Kaarsholm
N C
NC
Novo Research Institute, Novo Nordisk A/S, Bagsvaerd, Denmark.
Kolstrup
A M
AM
Danielsen
S E
SE
Holm
J
J
Hansen
S I
SI
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle; D002942:Circular Dichroism; D005260:Female; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008433:Mathematics; D008892:Milk / Q000378:metabolism; D008962:Models, Theoretical; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000032:analysis
D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D008024:Ligands; D011956:Receptors, Cell Surface; D014364:Tryptophan; D005492:Folic Acid