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Substrate Specificity, Inhibitor Selectivity and Structure-Function Relationships of Aldo-Keto Reductase 1B15: A Novel Human Retinaldehyde Reductase.

Giménez-Dejoz J, Kolář MH, Ruiz FX, Crespo I, Cousido-Siah A, Podjarny A, Barski OA, Fanfrlík J, Parés X, Farrés J, Porté S - PLoS ONE (2015)

Bottom Line: In contrast to AKR1B10, which strongly prefers all-trans-retinaldehyde, AKR1B15 exhibits superior catalytic efficiency with 9-cis-retinaldehyde, the best substrate found for this enzyme.Several typical AKR inhibitors do not significantly affect AKR1B15 activity.Amino acid substitutions clustered in loops A and C result in a smaller, more hydrophobic and more rigid active site in AKR1B15 compared with the AKR1B10 pocket, consistent with distinct substrate specificity and narrower inhibitor selectivity for AKR1B15.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Faculty of Biosciences, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain.

ABSTRACT
Human aldo-keto reductase 1B15 (AKR1B15) is a newly discovered enzyme which shares 92% amino acid sequence identity with AKR1B10. While AKR1B10 is a well characterized enzyme with high retinaldehyde reductase activity, involved in the development of several cancer types, the enzymatic activity and physiological role of AKR1B15 are still poorly known. Here, the purified recombinant enzyme has been subjected to substrate specificity characterization, kinetic analysis and inhibitor screening, combined with structural modeling. AKR1B15 is active towards a variety of carbonyl substrates, including retinoids, with lower kcat and Km values than AKR1B10. In contrast to AKR1B10, which strongly prefers all-trans-retinaldehyde, AKR1B15 exhibits superior catalytic efficiency with 9-cis-retinaldehyde, the best substrate found for this enzyme. With ketone and dicarbonyl substrates, AKR1B15 also shows higher catalytic activity than AKR1B10. Several typical AKR inhibitors do not significantly affect AKR1B15 activity. Amino acid substitutions clustered in loops A and C result in a smaller, more hydrophobic and more rigid active site in AKR1B15 compared with the AKR1B10 pocket, consistent with distinct substrate specificity and narrower inhibitor selectivity for AKR1B15.

No MeSH data available.


Related in: MedlinePlus

Comparison of the cofactor-binding site between the AKR1B15 model (A) and the AKR1B10 crystal structure (B).Interactions of Met265 and His269 with NADP+ in AKR1B15 are similar to those of Val265 and Arg269 in AKR1B10 (black dotted lines). His269 forms a π-stacking interaction with the adenine ring of the cofactor. The substitution of Lys22 by Arg in AKR1B15 prevents its interaction with the pyrophosphate bridge of NADP+. The salt bridge between Asp217 and Lys263 (red dotted line), acting as a safety belt in the coenzyme binding, and the π-stacking interaction of Tyr210 with the cofactor nicotinamide ring are conserved between the two AKRs. Carbon atoms of the cofactor are shown in green, whereas those of the enzyme are colored grey. Figures have been drawn using PyMOL.
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pone.0134506.g005: Comparison of the cofactor-binding site between the AKR1B15 model (A) and the AKR1B10 crystal structure (B).Interactions of Met265 and His269 with NADP+ in AKR1B15 are similar to those of Val265 and Arg269 in AKR1B10 (black dotted lines). His269 forms a π-stacking interaction with the adenine ring of the cofactor. The substitution of Lys22 by Arg in AKR1B15 prevents its interaction with the pyrophosphate bridge of NADP+. The salt bridge between Asp217 and Lys263 (red dotted line), acting as a safety belt in the coenzyme binding, and the π-stacking interaction of Tyr210 with the cofactor nicotinamide ring are conserved between the two AKRs. Carbon atoms of the cofactor are shown in green, whereas those of the enzyme are colored grey. Figures have been drawn using PyMOL.

Mentions: Regarding the cofactor-binding site, AKR1B15 shares with AKR1B10 all residues, except for Arg22, Met265 and His269 (Fig 5). The change of Lys22 in AKR1B10 to Arg in AKR1B15 may prevent its interaction with the pyrophosphate bridge of NADP+ but, due to the mobility of the Arg side chain in solution, its interaction with the cofactor cannot be excluded. Met265 and His269 keep interactions with the same groups of NADP+ that involve Val265 and Arg269 in AKR1B10 (i.e. a hydrogen bond between the N atom of Met265 and the 2′-phosphate group, a second hydrogen bond between the Nε of His269 and the 2′-phosphate group, and a stacking interaction of His269 with the adenine ring). Interestingly, a His residue at position 269 and its interactions have also been described in the rat AKR1B14 X-ray structure [53]. The salt bridge between the side-chain of Asp217 and Lys263, acting as a safety belt in cofactor binding, and the stacking interaction of Tyr210 side chain with the nicotinamide ring are also conserved. This is consistent with the AKR1B15 cofactor preference for NADP(H) and the low Km value of AKR1B15 for NADPH.


Substrate Specificity, Inhibitor Selectivity and Structure-Function Relationships of Aldo-Keto Reductase 1B15: A Novel Human Retinaldehyde Reductase.

Giménez-Dejoz J, Kolář MH, Ruiz FX, Crespo I, Cousido-Siah A, Podjarny A, Barski OA, Fanfrlík J, Parés X, Farrés J, Porté S - PLoS ONE (2015)

Comparison of the cofactor-binding site between the AKR1B15 model (A) and the AKR1B10 crystal structure (B).Interactions of Met265 and His269 with NADP+ in AKR1B15 are similar to those of Val265 and Arg269 in AKR1B10 (black dotted lines). His269 forms a π-stacking interaction with the adenine ring of the cofactor. The substitution of Lys22 by Arg in AKR1B15 prevents its interaction with the pyrophosphate bridge of NADP+. The salt bridge between Asp217 and Lys263 (red dotted line), acting as a safety belt in the coenzyme binding, and the π-stacking interaction of Tyr210 with the cofactor nicotinamide ring are conserved between the two AKRs. Carbon atoms of the cofactor are shown in green, whereas those of the enzyme are colored grey. Figures have been drawn using PyMOL.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519324&req=5

pone.0134506.g005: Comparison of the cofactor-binding site between the AKR1B15 model (A) and the AKR1B10 crystal structure (B).Interactions of Met265 and His269 with NADP+ in AKR1B15 are similar to those of Val265 and Arg269 in AKR1B10 (black dotted lines). His269 forms a π-stacking interaction with the adenine ring of the cofactor. The substitution of Lys22 by Arg in AKR1B15 prevents its interaction with the pyrophosphate bridge of NADP+. The salt bridge between Asp217 and Lys263 (red dotted line), acting as a safety belt in the coenzyme binding, and the π-stacking interaction of Tyr210 with the cofactor nicotinamide ring are conserved between the two AKRs. Carbon atoms of the cofactor are shown in green, whereas those of the enzyme are colored grey. Figures have been drawn using PyMOL.
Mentions: Regarding the cofactor-binding site, AKR1B15 shares with AKR1B10 all residues, except for Arg22, Met265 and His269 (Fig 5). The change of Lys22 in AKR1B10 to Arg in AKR1B15 may prevent its interaction with the pyrophosphate bridge of NADP+ but, due to the mobility of the Arg side chain in solution, its interaction with the cofactor cannot be excluded. Met265 and His269 keep interactions with the same groups of NADP+ that involve Val265 and Arg269 in AKR1B10 (i.e. a hydrogen bond between the N atom of Met265 and the 2′-phosphate group, a second hydrogen bond between the Nε of His269 and the 2′-phosphate group, and a stacking interaction of His269 with the adenine ring). Interestingly, a His residue at position 269 and its interactions have also been described in the rat AKR1B14 X-ray structure [53]. The salt bridge between the side-chain of Asp217 and Lys263, acting as a safety belt in cofactor binding, and the stacking interaction of Tyr210 side chain with the nicotinamide ring are also conserved. This is consistent with the AKR1B15 cofactor preference for NADP(H) and the low Km value of AKR1B15 for NADPH.

Bottom Line: In contrast to AKR1B10, which strongly prefers all-trans-retinaldehyde, AKR1B15 exhibits superior catalytic efficiency with 9-cis-retinaldehyde, the best substrate found for this enzyme.Several typical AKR inhibitors do not significantly affect AKR1B15 activity.Amino acid substitutions clustered in loops A and C result in a smaller, more hydrophobic and more rigid active site in AKR1B15 compared with the AKR1B10 pocket, consistent with distinct substrate specificity and narrower inhibitor selectivity for AKR1B15.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Faculty of Biosciences, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain.

ABSTRACT
Human aldo-keto reductase 1B15 (AKR1B15) is a newly discovered enzyme which shares 92% amino acid sequence identity with AKR1B10. While AKR1B10 is a well characterized enzyme with high retinaldehyde reductase activity, involved in the development of several cancer types, the enzymatic activity and physiological role of AKR1B15 are still poorly known. Here, the purified recombinant enzyme has been subjected to substrate specificity characterization, kinetic analysis and inhibitor screening, combined with structural modeling. AKR1B15 is active towards a variety of carbonyl substrates, including retinoids, with lower kcat and Km values than AKR1B10. In contrast to AKR1B10, which strongly prefers all-trans-retinaldehyde, AKR1B15 exhibits superior catalytic efficiency with 9-cis-retinaldehyde, the best substrate found for this enzyme. With ketone and dicarbonyl substrates, AKR1B15 also shows higher catalytic activity than AKR1B10. Several typical AKR inhibitors do not significantly affect AKR1B15 activity. Amino acid substitutions clustered in loops A and C result in a smaller, more hydrophobic and more rigid active site in AKR1B15 compared with the AKR1B10 pocket, consistent with distinct substrate specificity and narrower inhibitor selectivity for AKR1B15.

No MeSH data available.


Related in: MedlinePlus