SGC Structures

No. 549

ZADH1: Human zinc binding alcohol dehydrogenase, domain containing 1

Shafqat, N., Kavanagh, K.L., Pike, A.C.W., Pilka, E., Roos, A.K., Picaud, S., Johansson, C., Smee, C., Fedorov, O., Kochan, G., Gileadi, O., von Delft, F., Lee, W.H., Muller, S., Marsden, B.D., Bountra, C., Oppermann, U.

PDB Code: 2VNA

Datapack version: 1 (built on 3.Mar.08)

Description

Structure of human 15-oxoprostaglandin E2 D13 reductase type 2 (ZADH1): The product of the human ZADH1 (zinc containing alcohol dehydrogenase 1) gene has recently (1) been shown to catalyze the NADPH dependent reduction of 15-oxoprostaglandin E2 (15-oxo PGE2) to form 13,14-dihydro-15-oxo-PGE2. The PGE2 eicosanoid plays important roles in inflammation, and its inflammatory properties are attenuated by catabolism mediated by NADH dependent 15-hydroxyprostagladin dehydrogenase (HPGD) to form 15-oxo PGE2. This metabolite is further reduced by double bond reduction to yield 13,14-dihydro-15-oxo-PGE2, a reaction catalyzed by two distinct but related enzymes, namely LTB4Dh and ZADH1. The importance of HPGD and ZADH in mediating transcriptional activities of PPARγ was recently demonstrated, by being involved in production and inactivation of 15-oxo PGE2. This constitutes a novel mechanism involved in adipocyte differentiation through catabolism of PGE2 and regulation of ligand-induced PPARγ activation.

Whereas HPGD is a member of the short-chain dehydrogenases/reductases (SDR), noth LTB4DH and ZADH1 are members of the medium-chain-dehydrogenases/reductases (MDR), an evolutionarily conserved family of oxidoreductases. We previously determined the structures of human HPGD and LTB4DH, and we now provide the structure of human ZADH1. In contrast to the zinc containing alcohol dehydrogenases of the MDR family, ZADH1 and LTB4DH do not co tain metal ions.

MDR enzyme family: Medium-chain dehydrogenases/reductases (MDR) constitute a large enzyme superfamily with over 1000 members from all forms of life (2). The MDR enzymes represent many different activities of which the alcohol dehydrogenase system is studied in extensive detail (2). In mammals, MDR enzymes are besides the metabolism of alcohols involved in the detoxification of reactive carbonyls (aldehydes, ketones, quinones), and in the conversion of polyols, steroids and fatty acids or lipid mediators (2).

All MDR enzymes use NAD(H) and NADP(H) as cofactor and several but not all of the members require 1-2 zinc atoms for catalysis and structural stabilization as seen with the alcohol dehydrogenases. Based on this feature MDR enzymes are subgrouped into zinc-containing MDRs (containing alcohol dehydrogenases (ADH), polyol dehydrogenases (PDH), cinnamyl alcohol dehydrogenases (CAD) and yeast ADH), and non-zinc MDRs (containing leukotriene dehydrogenases (LTD), multifunctional respiratory enzymes (MRF), quinone oxidoreductases (QOR) and acyl CoA reductases (ACR) (2). Human ZADH1 belongs to the LTD subfamily of MDRs.

Structural features

Architecture of ZADH1: Structure determination of human ZADH1 (2VNA) in complex with NADPH reveals the two-domain α/β fold architecture of MDR enzymes. Typically these enzymes show a C-terminal coenzyme binding domain consisting of a Rossmann fold motif and an N-terminal domain for substrate binding. Surprisingly, the biological unit of human ZADH1 is a monomer since none of the crystallographically related subunits show any convincing oligomerization interaction as seen in other MDRs. The same observation is noted in the apostructure of the murine ortholog (PDB id 1vj1) (3). Superimposition of human ZADH1 with its closest structural neighbour, the murine ortholog reveals a nearly identical architecture with an rmsd of 1.9A. The largest difference is observed in the C-terminal cofactor binding domain, where the loop becomes ordered upon cofactor binding (Gly254-Pro266 in the human structure). The cofactor is bound in the cleft found between the two distinct domains.

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Cofactor binding in the human structure is achieved through several hydrogen bonding contacts comprising main-chain or side-chain interactions of residues Ala166, Cys167, Lys192, Tyr208, Cys253, Tyr259, Phe287, Val289 and Gly335. NADPH specificity is achieved by interactions of Lys192 with the 2’ phosphate group of the adenine ribose of the cofactor.

Human ZADH1 is related to another MDR enzyme, leukotriene B4 dehydrogenase (LTB4DH), with an overall sequence identity of about 40%. Superposition with the human paralog apostructure (rmsd 5.3 Å) reveals a similar domain arrangement and, although ordered in the LTB4DH apostructure, a displaced cosubstrate binding loop. Both enzymes catalyze the double bond reduction in eicosanoids, suggesting similar mechanisms and substrate specificities.
The reaction mechanism for ZADH1 double bond reductio based on the ternary complex of guinea pig LTB4DH and A. thaliana alkenal double bond reductase At5g16970 (4-5) is supposedly through formation of an α,β conjugated enolate intermediate, stabilized by interactions involving the substrate, the carbonyl-2-OH ribose, Tyr259 side-chain, and a water network. Hydride transfer occurs from the C4H of the R-side of the nicotinamide ring to the S-face of the electron deficient carbon center of the substrate, and subsequent protonation probably occurs directly from the solvent to the adjacent carbon of the substrate (4-5).

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References

1. Chou WL, Chuang LM, Chou CC, Wang AH, Lawson JA, FitzGerald GA, Chang ZF. Identification of a novel prostaglandin reductase reveals the involvement of prostaglandin E2 catabolism in regulation of peroxisome proliferator-activated receptor gamma activation. J Biol Chem. 2007; 282(25):18162-72.

2. Nordling E, Jornvall H, Persson B. Medium-chain dehydrogenases/reductases (MDR). Family characterizations including genome comparisons and active site modeling. Eur J Biochem. 2002;269(17):4267-76.

3. Levin, I., et al Crystal structure of a putative NADPH-dependent oxidoreductase (GI: 18204011) from mouse at 2.10 A resolution. Proteins v56 pp. 629-33, 2004

4. Youn B, Kim SJ, Moinuddin SG, Lee C, Bedgar DL, Harper AR, Davin LB, Lewis NG, Kang C. Mechanistic and structural studies of apoform, binary, and ternary complexes of the Arabidopsis alkenal double bond reductase At5g 6970. J Biol Chem. 2006;281(52):40076-88.

5. Hori T, Yokomizo T, Ago H, Sugahara M, Ueno G, Yamamoto M, Kumasaka T, Shimizu T, Miyano M. Structural basis of leukotriene B4 12- hydroxydehydrogenase/15-Oxo-prostaglandin 13-reductase catalytic mechanism and a possible Src homology 3 domain binding loop. J Biol Chem. 2004 May 21;279(21):22615-23.