NMR characterization of structure, backbone dynamics, and glutathione binding of the human macrophage migration inhibitory factor (MIF).
Human macrophage migration inhibitory factor is a 114 amino acid protein that belongs to the family of immunologic cytokines. Assignments of 1H, 15N, and 13C resonances have enabled the determination of the secondary structure of the protein, which consists of two alpha-helices (residues 18-31 and 89-72) and a central four-stranded beta-sheet. In the beta-sheet, two parallel beta-sheets are connected in an antiparallel sense. From the total of three cysteines present in the primary structure of MIF, none was found to form disulfide bridges. 1H-15N heteronuclear T1, T2, and steady-state NOE measurements indicate that the backbone of MIF exists in a rigid structure of limited conformational flexibility (on the nanosecond to picosecond time scale). Several residues located in the loop regions and at the N termini of two helices exhibit internal motions on the 1-3 ns time scale. The capacity to bind glutathione was investigated by titration of a uniform 15N-labeled sample and led us to conclude that MIF has, at best, very low affinity for glutathione.
Max Planck Institute for Biochemistry, Martinsried, Germany.
P
P
Mühlhahn
J
J
Bernhagen
M
M
Czisch
J
J
Georgescu
C
C
Renner
A
A
Ross
R
R
Bucala
T A
TA
Holak
D000595:Amino Acid Sequence; D001665:Binding Sites; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D008263:Macrophage Migration-Inhibitory Factors / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D008263:Macrophage Migration-Inhibitory Factors; D011993:Recombinant Fusion Proteins; D005978:Glutathione