TY - JOUR
T1 - The interrelationship between ligand binding and self-association of the folate binding protein
T2 - the role of detergent–tryptophan interaction
AU - Holm, Jan
AU - Schou, Christian
AU - Babol, Linnea N.
AU - Lawaetz, Anders Juul
AU - Bruun, Susanne Wrang
AU - Hansen, Morten Z.
AU - Hansen, Steen I.
PY - 2011/12
Y1 - 2011/12
N2 - Background: The folate binding protein (FBP) regulates homeostasis and intracellular trafficking of folic acid, a vitamin of decisive importance in cell division and growth. We analyzed whether interrelationship between ligand binding and self-association of FBP plays a significant role in the physiology of folate binding. Methods: Self-association behavior of apo- and holo-FBP was addressed through size exclusion chromatography, SDS-PAGE, mass spectrometry, surface plasmon resonance and fluorescence spectroscopy. Results: Especially holo-FBP exhibits concentration-dependent self-association at pH 7.4 (pI), and is more prone to associate into stable complexes than apo-FBP. Even more pronounced was the tendency to complexation between apo-FBP and holo-FBP in accord with a model predicting association between apo and holo monomers [19]. This will lead to removal of apo monomers from the reaction scheme resulting in a weak incomplete ligand binding similar to that observed at FBP concentrations < 10 nM. The presence of synthetic and natural detergents normalized folate binding kinetics and resulted in appearance of monomeric holo-FBP. Fluorescence spectroscopy indicated molecular interactions between detergent and tryptophan residues located in hydrophobic structures of apo-FBP which may participate in protein associations. General significance: Self-association into multimers may protect binding sites, and in case of holo-FBP even folate from biological degradation. High-affinity folate binding in body secretions, typically containing 1-10 nM FBP, requires the presence of natural detergents, i.e. cholesterol and phospholipids, to avoid complexation between apo- and holo-FBP.
AB - Background: The folate binding protein (FBP) regulates homeostasis and intracellular trafficking of folic acid, a vitamin of decisive importance in cell division and growth. We analyzed whether interrelationship between ligand binding and self-association of FBP plays a significant role in the physiology of folate binding. Methods: Self-association behavior of apo- and holo-FBP was addressed through size exclusion chromatography, SDS-PAGE, mass spectrometry, surface plasmon resonance and fluorescence spectroscopy. Results: Especially holo-FBP exhibits concentration-dependent self-association at pH 7.4 (pI), and is more prone to associate into stable complexes than apo-FBP. Even more pronounced was the tendency to complexation between apo-FBP and holo-FBP in accord with a model predicting association between apo and holo monomers [19]. This will lead to removal of apo monomers from the reaction scheme resulting in a weak incomplete ligand binding similar to that observed at FBP concentrations < 10 nM. The presence of synthetic and natural detergents normalized folate binding kinetics and resulted in appearance of monomeric holo-FBP. Fluorescence spectroscopy indicated molecular interactions between detergent and tryptophan residues located in hydrophobic structures of apo-FBP which may participate in protein associations. General significance: Self-association into multimers may protect binding sites, and in case of holo-FBP even folate from biological degradation. High-affinity folate binding in body secretions, typically containing 1-10 nM FBP, requires the presence of natural detergents, i.e. cholesterol and phospholipids, to avoid complexation between apo- and holo-FBP.
U2 - 10.1016/j.bbagen.2011.07.005
DO - 10.1016/j.bbagen.2011.07.005
M3 - Journal article
C2 - 21787841
SN - 0304-4165
VL - 1810
SP - 1330
EP - 1339
JO - B B A - General Subjects
JF - B B A - General Subjects
IS - 12
ER -