PGH2 Degradation Pathway Catalyzed by GSH-Heme Complex Bound Microsomal Prostaglandin E2 Synthase Type 2: The First Example of a Dual-Function Enzyme
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- 作者：Taro Yamada ; Fusao Takusagawa
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：July 17, 2007
- 年：2007
- 卷：46
- 期：28
- 页码：8414 - 8424
- 全文大小：548K
- 年卷期：v.46,no.28(July 17, 2007)
- ISSN：1520-4995
文摘
Prostaglandin E2 synthase (PGES) catalyzes the isomerization of PGH2 to PGE2. PGES type2 (mPGES-2) is a membrane-associated enzyme, whose N-terminal section is apparently inserted into thelipid bilayer. Both intact and N-terminal truncated enzymes have been isolated and have similar catalyticactivity. The recombinant N-terminal truncated enzyme purified from Escherichia coli HB101 grown inLB medium containing 
-aminolevulinate and Fe(NO3)3 has a red color, while the same enzyme purifiedfrom the same E. coli grown in minimal medium has no color. The red-colored enzyme has beencharacterized by mass, fluorescence, and EPR spectroscopies and X-ray crystallography. The enzyme isfound to contain bound glutathione (GSH) and heme. GSH binds to the active site with six H-bonds,while a heme is complexed with bound GSH forming a S-Fe coordination bond with no polar interactionwith mPGES-2. There is a large open space between the heme and the protein, where a PGH2 might beable to bind. The heme dissociation constant is 0.53 
M, indicating that mPGES-2 has relatively strongheme affinity. Indeed, expression of mPGES-2 in E. coli stimulates heme biosynthesis. Although mPGES-2has been reported to be a GSH-independent PGES, the crystal structure and sequence analysis indicatethat mPGES-2 is a GSH-binding protein. The GSH-heme complex-bound enzyme (mPGES-2h) catalyzesformation of 12(S)-hydroxy-5(Z),8(E),10(E)-heptadecatrienoic acid and malondialdehyde from PGH2, butnot formation of PGE2. The following kinetic parameters at 37 
C were determined: KM = 56 M, kcat= 63 s-1, and kcat/KM = 1.1 × 106 M-1 s-1. They suggest that mPGES-2h has significant catalytic activityfor PGH2 degradation. It is possible that both GSH-heme complex-free and -bound enzymes are presentin the same tissues. mPGES-2 in heme-rich liver is most likely to become the form of mPGES-2h andmight be involved in degradation reactions similar to that of cytochrome P450. Since mPGES-2 is anisomerase and mPGES-2h is a lyase, mPGES-2 cannot simply be classified into one of six classes set bythe International Union of Biochemistry and Molecular Biology.
-aminolevulinate and Fe(NO3)3 has a red color, while the same enzyme purifiedfrom the same E. coli grown in minimal medium has no color. The red-colored enzyme has beencharacterized by mass, fluorescence, and EPR spectroscopies and X-ray crystallography. The enzyme isfound to contain bound glutathione (GSH) and heme. GSH binds to the active site with six H-bonds,while a heme is complexed with bound GSH forming a S-Fe coordination bond with no polar interactionwith mPGES-2. There is a large open space between the heme and the protein, where a PGH2 might beable to bind. The heme dissociation constant is 0.53 M, indicating that mPGES-2 has relatively strongheme affinity. Indeed, expression of mPGES-2 in E. coli stimulates heme biosynthesis. Although mPGES-2has been reported to be a GSH-independent PGES, the crystal structure and sequence analysis indicatethat mPGES-2 is a GSH-binding protein. The GSH-heme complex-bound enzyme (mPGES-2h) catalyzesformation of 12(S)-hydroxy-5(Z),8(E),10(E)-heptadecatrienoic acid and malondialdehyde from PGH2, butnot formation of PGE2. The following kinetic parameters at 37 C were determined: KM = 56 M, kcat= 63 s-1, and kcat/KM = 1.1 × 106 M-1 s-1. They suggest that mPGES-2h has significant catalytic activityfor PGH2 degradation. It is possible that both GSH-heme complex-free and -bound enzymes are presentin the same tissues. mPGES-2 in heme-rich liver is most likely to become the form of mPGES-2h andmight be involved in degradation reactions similar to that of cytochrome P450. Since mPGES-2 is anisomerase and mPGES-2h is a lyase, mPGES-2 cannot simply be classified into one of six classes set bythe International Union of Biochemistry and Molecular Biology.