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Integration of Inhibition Kinetics and Molecular Dynamics Simulations: A Urea-Mediated Folding Study on Acetaldehyde Dehydrogenase 1

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Abstract

Understanding the mechanism of acetaldehyde dehydrogenase 1 (ALDH1) folding is important because this enzyme is directly involved in several types of cancers and other diseases. We investigated the urea-mediated unfolding of ALDH1 by integrating kinetic inhibition studies with computational molecular dynamics (MD) simulations. Conformational changes in the enzyme structure were also analyzed using intrinsic and 1-anilinonaphthalene-8-sulfonate (ANS)-binding fluorescence measurements. Kinetic studies revealed that the direct binding of urea to ALDH1 induces inactivation of ALDH1 in a manner of mixed-type inhibition. Tertiary structural changes associated with regional hydrophobic exposure of the active site were observed. The urea binding regions on ALDH1 were predicted by docking simulations and were partly shared with active site residues of ALDH1 and with interface residues of the oligomerization domain for tetramer formation. The docking results suggest that urea prevents formation of the ALDH1 normal shape for the tetramer state as well as entrance of the substrate into the active site. Our study provides insight into the structural changes that accompany urea-mediated unfolding of ALDH1 and the catalytic role associated with conformational changes.

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Abbreviations

ALDH1:

acetaldehyde dehydrogenase 1

ANS:

1-anilinonaphthalene-8-sulfonate

MD:

molecular dynamics

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Acknowledgments

Dr. Hang Mu was supported by a grant from the Science and Technology Planning Project of Jiaxing (No. 2013AY21032). Dr. Jinhyuk Lee was supported through grants from the KOBIC Research Support Program, KRIBB Research Initiative Program, and the Pioneer Research Center Program through the National Research Foundation of Korea funded through the Ministry of Science, ICT & Future Planning (2013M3C1A3064780). Dr. Yong-Doo Park was supported through a grant from the Zhejiang Provincial Natural Science Foundation of China, “Towards studying the function of C3dg protein and elucidating its role in the pathogenesis of atopic dermatitis” (Grant No. LY14H110001) and through a fund from the Science and Technology Planning Project of Jiaxing (No. 2014AY21026).

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    Authors and Affiliations

    1. Zhejiang Provincial Key Laboratory of Applied Enzymology, Yangtze Delta Region Institute of Tsinghua University, Jiaxing, 314006, People’s Republic of China

      Yingying Xu, Zhi-Rong Lü, Hang Mu & Yong-Doo Park

    2. School of Preclinical Medicine, Beijing University of Chinese Medicine, 11 Beisanhuan Dong Road, Beijing, 100029, People’s Republic of China

      Yingying Xu & Qian Zhang

    3. Korean Bioinformation Center (KOBIC), Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, South Korea

      Jinhyuk Lee

    4. Department of Nanobiotechnology and Bioinformatics, University of Sciences and Technology, Daejeon, 305-350, South Korea

      Jinhyuk Lee

    5. College of Biological and Environmental Sciences, Zhejiang Wanli University, Ningbo, 315100, People’s Republic of China

      Yong-Doo Park

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    1. Yingying Xu
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    Correspondence to Qian Zhang or Yong-Doo Park.

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    Yingying Xu and Jinhyuk Lee contributed equally to this work.

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    Xu, Y., Lee, J., Lü, ZR. et al. Integration of Inhibition Kinetics and Molecular Dynamics Simulations: A Urea-Mediated Folding Study on Acetaldehyde Dehydrogenase 1. Appl Biochem Biotechnol 179, 1101–1114 (2016). https://doi.org/10.1007/s12010-016-2052-5

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