Structures of human and porcine aldehyde reductase: an enzyme implicated in diabetic complications

120

AKR1D1 (steroid 5␤-reductase) reduces all ⌬ 4-3-ketosteroids to form 5␤-dihydrosteroids, a first step in the clearance of steroid hormones and an essential step in the synthesis of all bile acids. The reduction of the carbon-carbon double bond in an ␣,␤-unsaturated ketone by 5␤-reductase is a unique reaction in steroid enzymology because hydride transfer from NADPH to the ␤-face of a ⌬ 4 -3-ketosteroid yields a cis-A/B-ring configuration with an ϳ90°b end in steroid structure. Here, we report the first x-ray crystal structure of a mammalian steroid hormone carbon-carbon double bond reductase, human ⌬ 120. The Y58F and E120A mutants are devoid of activity, supporting a role for this dyad in the catalytic mechanism. Intriguingly, testosterone binds nonproductively, thereby rationalizing the substrate inhibition observed with this particular steroid. The locations of disease-linked mutations thought to be responsible for bile acid deficiency are also revealed.

mentioning

confidence: 99%

Crystal Structure of Human Liver Δ4-3-Ketosteroid 5β-Reductase (AKR1D1) and Implications for Substrate Binding and Catalysis

Costanzo

Drury

Penning

et al. 2008

120

“…The enzymatic properties of these invertebrate gene products have not been characterized yet; however, they were considered to be AKR superfamily enzymes because the catalytic tetrad (Asp-Tyr-Lys-His) known in AKR superfamily enzymes was well conserved in their sequences. Therefore, we chose the sequences of the Aplysia and Lottia putative enzymes (amino acid identity ϳ60% with HdRed) and those of well characterized human aldehyde reductase AKR1A1 (25)(26)(27)36) (identity 18% with HdRed) and a reductase from a halophilic bacterium Haloferax volcanii (AAB71807) (identity 36% with HdRed) for the comparison with HdRed. As shown in Fig.…”

Section: Fragments (P1 (Y(l/i)gasnm(l/i)-gwqmqr) P2 (D(l/i)vpsv(l/i)mentioning

confidence: 99%

A Novel Aldo-Keto Reductase, HdRed, from the Pacific Abalone Haliotis discus hannai, Which Reduces Alginate-derived 4-Deoxy-l-erythro-5-hexoseulose Uronic Acid to 2-Keto-3-deoxy-d-gluconate

Mochizuki

Nishiyama

Inoue

et al. 2015

Abalone feeds on brown seaweeds and digests seaweeds' alginate with alginate lyases (EC 4.2.2.3). However, it has been unclear whether the end product of alginate lyases (i.e. unsaturated monouronate-derived 4-deoxy-L-erythro-5-hexoseulose uronic acid (DEH)) is assimilated by abalone itself, because DEH cannot be metabolized via the Embden-Meyerhof pathway of animals. Under these circumstances, we recently noticed the occurrence of an NADPH-dependent reductase, which reduced DEH to 2-keto-3-deoxy-D-gluconate, in hepatopancreas extract of the pacific abalone Haliotis discus hannai. In the present study, we characterized this enzyme to some extent. The DEH reductase, named HdRed in the present study, could be purified from the acetone-dried powder of hepatopancreas by ammonium sulfate fractionation followed by conventional column chromatographies. HdRed showed a single band of ϳ40 kDa on SDS-PAGE and reduced DEH to 2-keto-3-deoxy-D-gluconate with an optimal temperature and pH at around 50°C and 7.0, respectively. HdRed exhibited no appreciable activity toward 28 authentic compounds, including aldehyde, aldose, ketose, ␣-keto-acid, uronic acid, deoxy sugar, sugar alcohol, carboxylic acid, ketone, and ester. The amino acid sequence of 371 residues of HdRed deduced from the cDNA showed 18 -60% identities to those of aldo-keto reductase (AKR) superfamily enzymes, such as human aldose reductase, halophilic bacterium reductase, and sea hare norsolorinic acid (a polyketide derivative) reductase-like protein. Catalytic residues and cofactor binding residues known in AKR superfamily enzymes were fairly well conserved in HdRed. Phylogenetic analysis for HdRed and AKR superfamily enzymes indicated that HdRed is an AKR belonging to a novel family.Alginate is a structural polysaccharide of brown seaweeds and certain bacteria, comprising ␤-D-mannuronate (M) 2 and ␣-L-guluronate (G), which form poly(M), poly(G), and random(MG) blocks in alginate polymer (1-3). Alginate from brown seaweeds has been widely used as a viscosifier and gelling agent in the food and pharmaceutical industries because sodium alginate solution exhibits high viscosity and forms an elastic gel upon forming calcium salt (1, 4). Alginate oligosaccharides are also recognized as functional materials that exhibit various biological functions, such as promotion of root growth of higher plants (5, 6), acceleration of growth rate of Bifidobacterium sp. (7), and stimulation of proliferation of endothelial cells (8). Further, 4-deoxy-5-erythro-hexoseulose uronic acid (DEH), the end product of alginate lyases (EC 4.2.2.3 and EC 4.2.2.11) (see Fig. 1), was recently proven to be available for ethanol fermentation with genetically modified microorganisms (9 -11). These findings have increased the practical potentiality of both alginate and alginate-producing brown seaweeds. Besides such practical aspects, alginate is an important food source for herbivorous gastropods like abalone (12-16). Namely, abalone feeds brown seaweeds as a daily diet and digests poly(M) block of ...

“…The aldo-keto reductases are a superfamily of enzymes that share a number of common structural features; in particular they have a (␤/␣) 8 -barrel structure that was first demonstrated for human aldose reductase (AKR1B1) (6,7). Subsequently the crystal structures of seven other members of the aldo-keto reductase family have been determined: porcine aldehyde reductase (AKR1A1) (8), rat liver 3␣-hydroxysteroid dehydrogenase (AKR1C1) (9), mouse FR1 protein (AKR1B8) (10), bacterial 2,5-diketo-D-gluconic acid reductase (AKR5C) (11), Chinese hamster ovary reductase (AKR1B9) (12), and recently yeast GCY1 (AKR3A1) (13) and human 3␣-hydroxysteroid dehydrogenase (AKR1C2) (14).…”

Section: Was Purified As a Cytosolic Nad(p)mentioning

confidence: 99%

“…Subsequently the crystal structures of seven other members of the aldo-keto reductase family have been determined: porcine aldehyde reductase (AKR1A1) (8), rat liver 3␣-hydroxysteroid dehydrogenase (AKR1C1) (9), mouse FR1 protein (AKR1B8) (10), bacterial 2,5-diketo-D-gluconic acid reductase (AKR5C) (11), Chinese hamster ovary reductase (AKR1B9) (12), and recently yeast GCY1 (AKR3A1) (13) and human 3␣-hydroxysteroid dehydrogenase (AKR1C2) (14). These enzymes show considerable diversity in terms of substrate specificity, catalyzing the reduction of a range of aliphatic and aromatic aldehydes and ketones, including sugars, steroids, and xenobiotics.…”

Section: Was Purified As a Cytosolic Nad(p)mentioning

confidence: 99%

The Crystal Structure of Rat Liver AKR7A1

Kozma

Brown

Ellis

et al. 2002

The structure of the rat liver aflatoxin dialdehyde reductase (AKR7A1) has been solved to 1.38-Å resolution. Although it shares a similar ␣/␤-barrel structure with other members of the aldo-keto reductase superfamily, AKR7A1 is the first dimeric member to be crystallized. The crystal structure also reveals details of the ternary complex as one subunit of the dimer contains NADP ؉ and the inhibitor citrate. Although the underlying catalytic mechanism appears similar to other aldoketo reductases, the substrate-binding pocket contains several charged amino acids (Arg-231 and Arg-327) that distinguish it from previously characterized aldo-keto reductases with respect to size and charge. These differences account for the substrate specificity for 4-carbon acid-aldehydes such as succinic semialdehyde and 2-carboxybenzaldehyde as well as for the idiosyncratic substrate aflatoxin B 1 dialdehyde of this subfamily of enzymes. Structural differences between the AKR7A1 ternary complex and apoenzyme reveal a significant hinged movement of the enzyme involving not only the loops of the structure but also parts of the ␣/␤-barrel most intimately involved in cofactor binding.Rat liver aflatoxin dialdehyde reductase (AFAR; AKR7A1) 1 was purified as a cytosolic NAD(P) ϩ -dependent reductase that is capable of reducing the dialdehyde phenolate metabolite of aflatoxin B 1 (1, 2). The enzyme is of particular importance as it contributes toward resistance to the hepatocarcinogen aflatoxin B 1 in rats (2) and is inducible in rat liver through the inclusion of either natural or synthetic compounds in the diet (1, 3, 4). Cloning and sequencing of the gene encoding this enzyme (3) identified it as belonging to the aldo-keto reductase (AKR) superfamily (5) and led to its assignation as the founder member of the AKR7 family (5).The aldo-keto reductases are a superfamily of enzymes that share a number of common structural features; in particular they have a (␤/␣) 8 -barrel structure that was first demonstrated for human aldose reductase (AKR1B1) (6, 7). Subsequently the crystal structures of seven other members of the aldo-keto reductase family have been determined: porcine aldehyde reductase (AKR1A1) (8), rat liver 3␣-hydroxysteroid dehydrogenase (AKR1C1) (9), mouse FR1 protein (AKR1B8) (10), bacterial 2,5-diketo-D-gluconic acid reductase (AKR5C) (11), Chinese hamster ovary reductase (AKR1B9) (12), and recently yeast GCY1 (AKR3A1) (13) and human 3␣-hydroxysteroid dehydrogenase (AKR1C2) (14). These enzymes show considerable diversity in terms of substrate specificity, catalyzing the reduction of a range of aliphatic and aromatic aldehydes and ketones, including sugars, steroids, and xenobiotics. Although structural information has revealed significant differences in the substrate-binding pocket, the relationships between structure and substrate specificity have not been fully determined. Site-directed mutagenesis of residues predicted to have important roles in binding of substrate and in catalysis has provided limited information on substra...