Crystal Structure of Macrophage Migration Inhibitory Factor Complexed with (E)-2-Fluoro-p-hydroxycinnamate at 1.8 Å Resolution:  Implications for Enzymatic Catalysis and Inhibition

Macrophage migration inhibitory factor (MIF) exhibits dual activities. It acts as an immunoregulatory protein as well as a phenylpyruvate tautomerase. To understand better the relationship between these two activities and to elucidate the structural basis for the enzymatic activity, a crystal struct...

Full description

Saved in:
Bibliographic Details
Published in:Biochemistry (Easton) 1999-06, Vol.38 (23), p.7444-7452
Main Authors: Taylor, Alexander B, Johnson, William H, Czerwinski, Robert M, Li, Horng-Shin, Hackert, Marvin L, Whitman, Christian P
Format: Article
Language:English
Subjects:
Animals
Binding, Competitive
Catalysis
Cinnamates - chemistry
Cinnamates - metabolism
Coumaric Acids - chemistry
Coumaric Acids - metabolism
Crystallization
Crystallography, X-Ray
Enzyme Inhibitors - chemistry
Enzyme Inhibitors - metabolism
Fatty Acids, Unsaturated - chemistry
Intramolecular Oxidoreductases - antagonists & inhibitors
Intramolecular Oxidoreductases - chemistry
Intramolecular Oxidoreductases - metabolism
Isomerases - antagonists & inhibitors
Kinetics
Macromolecular Substances
Macrophage Migration-Inhibitory Factors - antagonists & inhibitors
Macrophage Migration-Inhibitory Factors - chemistry
Macrophage Migration-Inhibitory Factors - metabolism
Mice
Models, Molecular
Protein Binding
Sequence Homology, Amino Acid
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Macrophage migration inhibitory factor (MIF) exhibits dual activities. It acts as an immunoregulatory protein as well as a phenylpyruvate tautomerase. To understand better the relationship between these two activities and to elucidate the structural basis for the enzymatic activity, a crystal structure of a complex between murine MIF and (E)-2-fluoro-p-hydroxycinnamate, a competitive inhibitor of the tautomerase activity, has been determined to 1.8 Å resolution. The structure is nearly superimposable on that of the free protein indicating that the presence of the inhibitor does not result in any major structural changes. The inhibitor also confirms the location of the active site in a hydrophobic cavity containing the amino-terminal proline. Within this cavity, the inhibitor interacts with residues from adjacent subunits. At the back of the cavity, the side-chain carbonyl oxygen of Asn-97‘ interacts with the phenolic hydroxyl group of the inhibitor while at the mouth of the cavity the ammonium group of Lys-32 interacts with a carboxylate oxygen. The other carboxylate oxygen of the inhibitor interacts with Pro-1. The hydroxyl group of Tyr-95‘ interacts weakly with the fluoro group on the inhibitor. The hydrophobic side chains of five active-site residues (Met-2, Ile-64, Met-101, Val-106, and Phe-113) and the phenyl moiety of Tyr-95‘ are responsible for the binding of the phenyl group. Further insight into the enzymatic activity of MIF was obtained by carrying out kinetic studies using the enol isomers of phenylpyruvate and (p-hydroxyphenyl)pyruvate. The results demonstrate that MIF processes the enol isomers more efficiently than the keto isomers primarily because of a decrease in K m. On the basis of these results, a mechanism is proposed for the MIF-catalyzed tautomerization reaction.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi9904048