Structural basis of leukotriene B4 12-hydroxydehydrogenase/15-Oxo-prostaglandin 13-reductase catalytic mechanism and a possible Src homology 3 domain binding loop.
The bifunctional leukotriene B(4) 12-hydroxydehydrogenase/15-oxo-prostaglandin 13-reductase (LTB(4) 12-HD/PGR) is an essential enzyme for eicosanoid inactivation. It is involved in the metabolism of the E and F series of 15-oxo-prostaglandins (15-oxo-PGs), leukotriene B(4) (LTB(4)), and 15-oxo-lipoxin A(4) (15-oxo-LXA(4)). Some nonsteroidal anti-inflammatory drugs (NSAIDs), which primarily act as cyclooxygenase inhibitors also inhibit LTB(4) 12-HD/PGR activity. Here we report the crystal structure of the LTB(4) 12-HD/PGR, the binary complex structure with NADP(+), and the ternary complex structure with NADP(+) and 15-oxo-PGE(2). In the ternary complex, both in the crystalline form and in solution, the enolate anion intermediate accumulates as a brown chromophore. PGE(2) contains two chains, but only the omega-chain of 15-oxo-PGE(2) was defined in the electron density map in the ternary complex structure. The omega-chain was identified at the hydrophobic pore on the dimer interface. The structure showed that the 15-oxo group forms hydrogen bonds with the 2'-hydroxyl group of nicotine amide ribose of NADP(+) and a bound water molecule to stabilize the enolate intermediate during the reductase reaction. The electron-deficient C13 atom of the conjugated enolate may be directly attacked by a hydride from the NADPH nicotine amide in a stereospecific manner. The moderate recognition of 15-oxo-PGE(2) is consistent with a broad substrate specificity of LTB(4) 12-HD/PGR. The structure also implies that a Src homology domain 3 may interact with the left-handed proline-rich helix at the dimer interface and regulate LTB(4) 12-HD/PGR activity by disruption of the substrate binding pore to accommodate the omega-chain.
Structural Biophysics Laboratory, Highthroughput Factory, Coherent X-ray Optics Laboratory, RIKEN Harima Institute at SPring-8, 1-1-1 Kouto, Mikazuki, Sayo-gun, Hyogo 679-5148, Japan.
Tetsuya
T
Hori
Takehiko
T
Yokomizo
Hideo
H
Ago
Mitsuaki
M
Sugahara
Go
G
Ueno
Masaki
M
Yamamoto
Takashi
T
Kumasaka
Takao
T
Shimizu
Masashi
M
Miyano
D011452:15-Oxoprostaglandin 13-Reductase / Q000737:chemistry*; D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D002384:Catalysis; D002712:Chlorides / Q000737:chemistry; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D019281:Dimerization; D015777:Eicosanoids / Q000737:chemistry; D004583:Electrons; D006168:Guinea Pigs; D006860:Hydrogen Bonding; D007477:Ions; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009249:NADP / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D013379:Substrate Specificity; D014867:Water / Q000737:chemistry; D014961:X-Ray Diffraction; D018909:src Homology Domains
D000894:Anti-Inflammatory Agents, Non-Steroidal; D002712:Chlorides; D016861:Cyclooxygenase Inhibitors; D015777:Eicosanoids; D007477:Ions; D014867:Water; D009249:NADP; D000429:Alcohol Oxidoreductases; C082646:leukotriene B4 12-hydroxydehydrogenase; D011452:15-Oxoprostaglandin 13-Reductase