Binding of biliverdin, bilirubin, and thyroid hormones to lipocalin-type prostaglandin D synthase.
Lipocalin-type prostaglandin D synthase is a major protein of the cerebrospinal fluid and was originally known as beta-trace. We investigated the binding ability of prostaglandin D synthase toward bile pigments, thyroid hormones, steroid hormones, and fatty acids in this present study. We found that the recombinant enzyme binds bile pigments and thyroid hormones, resulting in quenching of the intrinsic tryptophan fluorescence, the appearance of induced circular dichroism of the lipophilic ligands, and a red shift of the absorption spectra of bilirubin and biliverdin. The binding of prostaglandin D synthase to lipophilic ligands was also demonstrated by the resonant mirror technique and surface plasmon resonance detection. The dissociation constants were calculated to be 33 nM, 37 nM, 660 nM, 820 nM, and 2.08 microM for biliverdin, bilirubin, L-thyroxine, 3,3',5'-triiodo-L-thyronine, and 3,3', 5-triiodo-L-thyronine, respectively. Biliverdin and bilirubin underwent a shift in their absorption peaks from 375 to 380 nm and from 439 to 446 nm, respectively, after binding to prostaglandin D synthase. Bilirubin bound to the enzyme showed a bisignate CD spectrum with a (-) Cotton effect at 422 nm and a (+) Cotton effect at 472 nm, indicating a right-handed chirality. The ligands also inhibited prostaglandin D synthase activity noncompetitively in a concentration-dependent manner, with IC50 values between 3.9 and 10. 9 microM. Epididymal retinoic acid-binding protein and beta-lactoglobulin, two other lipocalin proteins that bind retinoids such as prostaglandin D synthase, did not show any significant interaction with bile pigments or thyroid hormones. These results show that prostaglandin D synthase binds small lipophilic ligands with a specificity distinct from that of other lipocalins.
Beuckmann
C T
CT
Osaka Bioscience Institute, Suita, Japan.
Aoyagi
M
M
Okazaki
I
I
Hiroike
T
T
Toh
H
H
Hayaishi
O
O
Urade
Y
Y
D000818:Animals; D001609:Beta-Globulins / Q000737:chemistry / Q000378:metabolism; D001663:Bilirubin / Q000737:chemistry* / Q000378:metabolism; D001664:Biliverdine / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000737:chemistry; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D019746:Intramolecular Oxidoreductases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D007782:Lactoglobulins / Q000737:chemistry; D008024:Ligands; D054834:Lipocalins; D008958:Models, Molecular; D018996:Myelin P2 Protein / Q000737:chemistry / Q000378:metabolism; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D051381:Rats; D018168:Receptors, Retinoic Acid / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013963:Thyroid Hormones / Q000737:chemistry* / Q000378:metabolism
D001609:Beta-Globulins; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D007782:Lactoglobulins; D008024:Ligands; D054834:Lipocalins; D018996:Myelin P2 Protein; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D018168:Receptors, Retinoic Acid; D013963:Thyroid Hormones; D019746:Intramolecular Oxidoreductases; C022466:prostaglandin R2 D-isomerase; D001664:Biliverdine; D001663:Bilirubin