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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

Molecular docking of substrates or inhibitors to the active-site pocket of AKR1B15.(A) Residues implicated in binding all-trans- and 9-cis-retinaldehyde are displayed in light and dark grey sticks; while the substrates are shown in light and dark blue, respectively. The residues found in the most external part of all-trans-retinaldehyde binding channel in AKR1B10 are highlighted in yellow. The energy minimized apo-conformation is displayed in magenta cartoon. (B) and (C) Side view of the surface contour of the active-site pocket, depicted in grey and orange for AKR1B15 and AKR1B10, respectively, to show the inhibitor “specificity pocket”. A thick grey curved line indicates the “specificity pocket” in AKR1B10. As it is shown, this pocket may not be opened in AKR1B15, likely due to the presence of bulky Phe residues. (D) The inhibitor JF0064 (PDB ID 4ICC) bound to AKR1B15 is displayed as sticks with C atoms in magenta, while residues interacting with the inhibitor are shown as sticks with C atoms in grey. (E) Steric hindrance preventing tolrestat (in blue) and sorbinil (in orange) from binding to the active site of AKR1B15. For this analysis, the AKR1B15 structure model was superimposed with the AKR1B10 crystallographic structures with tolrestat (PDB ID 1ZUA) and sorbinil (PDB ID 4GA8). NADP+ is colored in orange. Figures have been drawn using PyMOL.
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pone.0134506.g006: Molecular docking of substrates or inhibitors to the active-site pocket of AKR1B15.(A) Residues implicated in binding all-trans- and 9-cis-retinaldehyde are displayed in light and dark grey sticks; while the substrates are shown in light and dark blue, respectively. The residues found in the most external part of all-trans-retinaldehyde binding channel in AKR1B10 are highlighted in yellow. The energy minimized apo-conformation is displayed in magenta cartoon. (B) and (C) Side view of the surface contour of the active-site pocket, depicted in grey and orange for AKR1B15 and AKR1B10, respectively, to show the inhibitor “specificity pocket”. A thick grey curved line indicates the “specificity pocket” in AKR1B10. As it is shown, this pocket may not be opened in AKR1B15, likely due to the presence of bulky Phe residues. (D) The inhibitor JF0064 (PDB ID 4ICC) bound to AKR1B15 is displayed as sticks with C atoms in magenta, while residues interacting with the inhibitor are shown as sticks with C atoms in grey. (E) Steric hindrance preventing tolrestat (in blue) and sorbinil (in orange) from binding to the active site of AKR1B15. For this analysis, the AKR1B15 structure model was superimposed with the AKR1B10 crystallographic structures with tolrestat (PDB ID 1ZUA) and sorbinil (PDB ID 4GA8). NADP+ is colored in orange. Figures have been drawn using PyMOL.

Mentions: As it has been described above, AKR1B15 is active towards retinoids, and thus the binding mode of all-trans- and 9-cis-retinaldehyde was analyzed. The obtained models were also analyzed with the QMEAN server and displayed similar scores (0.68/1 and 0.69/1, respectively) as the AKR1B15 holoenzyme model. The analysis showed that both substrates could be placed with their carbonyl groups at catalytic distance from the hydroxyl group of Tyr49, the Nε of His111, and the cofactor C4 atom (2.9, 2.9, and 3.2 Å, respectively). The two molecules would be positioned in a similar manner into a narrow and hydrophobic pocket, establishing contacts with Trp21, Phe48, Phe123, Trp220, and Phe301 (all-trans-retinaldehyde), and Phe48, Trp220, Phe299, and Phe301 (9-cis-retinaldehyde) (Fig 6A). A slight rearrangement of loop A and loop C (Fig 6A) could allow the establishment of a hydrogen-bond or an electrostatic interaction between Lys125 and Glu303 (Fig 6B).


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)

Molecular docking of substrates or inhibitors to the active-site pocket of AKR1B15.(A) Residues implicated in binding all-trans- and 9-cis-retinaldehyde are displayed in light and dark grey sticks; while the substrates are shown in light and dark blue, respectively. The residues found in the most external part of all-trans-retinaldehyde binding channel in AKR1B10 are highlighted in yellow. The energy minimized apo-conformation is displayed in magenta cartoon. (B) and (C) Side view of the surface contour of the active-site pocket, depicted in grey and orange for AKR1B15 and AKR1B10, respectively, to show the inhibitor “specificity pocket”. A thick grey curved line indicates the “specificity pocket” in AKR1B10. As it is shown, this pocket may not be opened in AKR1B15, likely due to the presence of bulky Phe residues. (D) The inhibitor JF0064 (PDB ID 4ICC) bound to AKR1B15 is displayed as sticks with C atoms in magenta, while residues interacting with the inhibitor are shown as sticks with C atoms in grey. (E) Steric hindrance preventing tolrestat (in blue) and sorbinil (in orange) from binding to the active site of AKR1B15. For this analysis, the AKR1B15 structure model was superimposed with the AKR1B10 crystallographic structures with tolrestat (PDB ID 1ZUA) and sorbinil (PDB ID 4GA8). NADP+ is colored in orange. Figures have been drawn using PyMOL.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0134506.g006: Molecular docking of substrates or inhibitors to the active-site pocket of AKR1B15.(A) Residues implicated in binding all-trans- and 9-cis-retinaldehyde are displayed in light and dark grey sticks; while the substrates are shown in light and dark blue, respectively. The residues found in the most external part of all-trans-retinaldehyde binding channel in AKR1B10 are highlighted in yellow. The energy minimized apo-conformation is displayed in magenta cartoon. (B) and (C) Side view of the surface contour of the active-site pocket, depicted in grey and orange for AKR1B15 and AKR1B10, respectively, to show the inhibitor “specificity pocket”. A thick grey curved line indicates the “specificity pocket” in AKR1B10. As it is shown, this pocket may not be opened in AKR1B15, likely due to the presence of bulky Phe residues. (D) The inhibitor JF0064 (PDB ID 4ICC) bound to AKR1B15 is displayed as sticks with C atoms in magenta, while residues interacting with the inhibitor are shown as sticks with C atoms in grey. (E) Steric hindrance preventing tolrestat (in blue) and sorbinil (in orange) from binding to the active site of AKR1B15. For this analysis, the AKR1B15 structure model was superimposed with the AKR1B10 crystallographic structures with tolrestat (PDB ID 1ZUA) and sorbinil (PDB ID 4GA8). NADP+ is colored in orange. Figures have been drawn using PyMOL.
Mentions: As it has been described above, AKR1B15 is active towards retinoids, and thus the binding mode of all-trans- and 9-cis-retinaldehyde was analyzed. The obtained models were also analyzed with the QMEAN server and displayed similar scores (0.68/1 and 0.69/1, respectively) as the AKR1B15 holoenzyme model. The analysis showed that both substrates could be placed with their carbonyl groups at catalytic distance from the hydroxyl group of Tyr49, the Nε of His111, and the cofactor C4 atom (2.9, 2.9, and 3.2 Å, respectively). The two molecules would be positioned in a similar manner into a narrow and hydrophobic pocket, establishing contacts with Trp21, Phe48, Phe123, Trp220, and Phe301 (all-trans-retinaldehyde), and Phe48, Trp220, Phe299, and Phe301 (9-cis-retinaldehyde) (Fig 6A). A slight rearrangement of loop A and loop C (Fig 6A) could allow the establishment of a hydrogen-bond or an electrostatic interaction between Lys125 and Glu303 (Fig 6B).

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