Structure of the tetrameric form of human L‐Xylulose reductase: Probing the inhibitor‐binding site with molecular modeling and site‐directed mutagenesis

Abstract

L-Xylulose reductase (XR) is a member of the short-chain dehydrogenase/reductase (SDR) superfamily. In this study we report the structure of the biological tetramer of human XR in complex with NADP(+) and a competitive inhibitor solved at 2.3 A resolution. A single subunit of human XR is formed by a centrally positioned, seven-stranded, parallel beta-sheet surrounded on either side by two arrays of three alpha-helices. Two helices located away from the main body of the protein form the variable substrate-binding cleft, while the dinucleotide coenzyme-binding motif is formed by a classical Rossmann fold. The tetrameric structure of XR, which is held together via salt bridges formed by the guanidino group of Arg203 from one monomer and the carboxylate group of the C-terminal residue Cys244 from the neighboring monomer, explains the ability of human XR to prevent the cold inactivation seen in the rodent forms of the enzyme. The orientations of Arg203 and Cys244 are maintained by a network of hydrogen bonds and main-chain interactions of Gln137, Glu238, Phe241, and Trp242. These interactions are similar to those defining the quaternary structure of the closely related carbonyl reductase from mouse lung. Molecular modeling and site-directed mutagenesis identified the active site residues His146 and Trp191 as forming essential contacts with inhibitors of XR. These results could provide a structural basis in the design of potent and specific inhibitors for human XR.

Garrod's fourth inborn error of metabolism solved by the identification of mutations causing pentosuria

“…DCXR functions in vivo as a homotetramer (13), requiring the dinucleotide cofactor NADPH. Examination of the 3D structure of DCXR bound to NADP + indicates that residues in the Cterminal region are important for interaction between individual monomers (14,18). Given that the protein encoded by the DCXR c.583ΔC allele lacks more than 50 C-terminal amino acids of the 244-aa protein, formation of a functional tetramer by this allele is likely to be reduced or eliminated.…”

“…3) are also likely to lead to loss of normal protein function. The deleted region of the first mutant transcript from DCXR c.52(+1)G > A encodes a large part of the NADP + cofactor-binding domain (14,18). The second mutant transcript from DCXR c.52(+1)G > A encodes only 15 wild-type amino acids.…”

Transgenic Mice Over-expressing Dicarbonyl/L-xylulose Reductase Gene Crossed with KK-Ay Diabetic Model Mice: An Animal Model for the Metabolism of Renal Carbonyl Compounds

“…The enzyme belongs to the same short chain carbonyl reductase subfamily as the murine lung-specific carbonyl reductase which shares 63% identity at the amino acid sequence level [12]. The crystallographic analysis of DCXR was recently achieved [1,2].…”

Suppression of AGE Precursor Formation Following Unilateral Ureteral Obstruction in Mouse Kidneys by Transgenic Expression of α-Dicarbonyl/L-Xylulose Reductase

“…13,14) To examine the relevance of DCXR in the kidney, we generated transgenic (Tg) mice for DCXR. 15) Further crossing the DCXR Tg mice with KK-A y type 2 diabetic model mice, we found that over-expression of DCXR suppressed the increase in body weight and serum glucose levels associated with a diabetes-prone genetic background in the hybrid Tg/A y mice as compared to their +/A y littermates.…”