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Identification of important residues in diketoreductase from Acinetobacter baylyi by molecular modeling and site-directed mutagenesis.
Biochimie. 2012 Feb; 94(2):471-8.B

Abstract

Diketoreductase (DKR) from Acinetobacter baylyi exhibits a unique property of double reduction of a β, δ-diketo ester with excellent stereoselectivity, which can serve as an efficient biocatalyst for the preparation of an important chiral intermediate for cholesterol lowering statin drugs. Taken the advantage of high homology between DKR and human heart 3-hydroxyacyl-CoA dehydrogenase (HAD), a molecular model was created to compare the tertiary structures of DKR and HAD. In addition to the possible participation of His-143 in the enzyme catalysis by pH profile, three key amino acid residues, Ser-122, His-143 and Glu-155, were identified and mutated to explore the possibility of involving in the catalytic process. The catalytic activities for mutants S122A/C, H143A/K and E155Q were below detectable level, while their binding affinities to the diketo ester substrate and cofactor NADH did not change obviously. The experimental results were further supported by molecular docking, suggesting that Ser-122 and His-143 were essential for the proton transfer to the carbonyl functional groups of the substrate. Moreover, Glu-155 was crucial for maintaining the proper orientation and protonation of the imidazole ring of His-143 for efficient catalysis.

Authors+Show Affiliations

Laboratory of Chemical Biology, China Pharmaceutical University, 24 Tongjia Street, Nanjing, Jiangsu Province 210009, People's Republic of China.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

21893158

Citation

Huang, Yan, et al. "Identification of Important Residues in Diketoreductase From Acinetobacter Baylyi By Molecular Modeling and Site-directed Mutagenesis." Biochimie, vol. 94, no. 2, 2012, pp. 471-8.
Huang Y, Lu Z, Liu N, et al. Identification of important residues in diketoreductase from Acinetobacter baylyi by molecular modeling and site-directed mutagenesis. Biochimie. 2012;94(2):471-8.
Huang, Y., Lu, Z., Liu, N., & Chen, Y. (2012). Identification of important residues in diketoreductase from Acinetobacter baylyi by molecular modeling and site-directed mutagenesis. Biochimie, 94(2), 471-8. https://doi.org/10.1016/j.biochi.2011.08.015
Huang Y, et al. Identification of Important Residues in Diketoreductase From Acinetobacter Baylyi By Molecular Modeling and Site-directed Mutagenesis. Biochimie. 2012;94(2):471-8. PubMed PMID: 21893158.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of important residues in diketoreductase from Acinetobacter baylyi by molecular modeling and site-directed mutagenesis. AU - Huang,Yan, AU - Lu,Zhuo, AU - Liu,Nan, AU - Chen,Yijun, Y1 - 2011/08/30/ PY - 2011/06/06/received PY - 2011/08/23/accepted PY - 2011/9/7/entrez PY - 2011/9/7/pubmed PY - 2012/6/12/medline SP - 471 EP - 8 JF - Biochimie JO - Biochimie VL - 94 IS - 2 N2 - Diketoreductase (DKR) from Acinetobacter baylyi exhibits a unique property of double reduction of a β, δ-diketo ester with excellent stereoselectivity, which can serve as an efficient biocatalyst for the preparation of an important chiral intermediate for cholesterol lowering statin drugs. Taken the advantage of high homology between DKR and human heart 3-hydroxyacyl-CoA dehydrogenase (HAD), a molecular model was created to compare the tertiary structures of DKR and HAD. In addition to the possible participation of His-143 in the enzyme catalysis by pH profile, three key amino acid residues, Ser-122, His-143 and Glu-155, were identified and mutated to explore the possibility of involving in the catalytic process. The catalytic activities for mutants S122A/C, H143A/K and E155Q were below detectable level, while their binding affinities to the diketo ester substrate and cofactor NADH did not change obviously. The experimental results were further supported by molecular docking, suggesting that Ser-122 and His-143 were essential for the proton transfer to the carbonyl functional groups of the substrate. Moreover, Glu-155 was crucial for maintaining the proper orientation and protonation of the imidazole ring of His-143 for efficient catalysis. SN - 1638-6183 UR - https://www.unboundmedicine.com/medline/citation/21893158/Identification_of_important_residues_in_diketoreductase_from_Acinetobacter_baylyi_by_molecular_modeling_and_site_directed_mutagenesis_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0300-9084(11)00330-0 DB - PRIME DP - Unbound Medicine ER -