Abstract

The bifunctional enzyme Δ1-pyrroline-5-carboxylate synthase (P5CS) is vital to the synthesis of proline and ornithine, playing an essential role in human health and agriculture. Pathogenic mutations in P5CS gene (ALDH18A1) lead to neurocutaneous syndrome and skin relaxation connective tissue disease in humans, and P5CS deficiency seriously damages the ability to resist adversity in plants. We have recently found that P5CS forms the cytoophidium in vivo and filaments in vitro. However, it is difficult to appreciate the function of P5CS filamentation without precise structures. Using cryo-electron microscopy, here we solve structures of Drosophila full-length P5CS in three states at resolution from 3.1 to 4.3 Å. We observe distinct ligand-binding states and conformational changes for the GK and GPR domains, respectively. Divergent helical filaments are assembled by P5CS tetramers and stabilized by multiple interfaces. Point mutations disturbing those interfaces prevent P5CS filamentation and greatly reduce the enzymatic activity. Our findings reveal that filamentation is crucial for the coordination between the GK and GPR domains, providing structural basis for catalytic function of P5CS filaments.

Data availability

7F5T 7F5U 7F5V 7F5X 7WX3 7WX4 7WXF 7WXG 7WXH 7WXIFigure 5-Source Data 1 in enzyme activity assay, which related to Figure 5D;Validation summary report as Related Manuscript File;Maps and coordinates data as a Supporting Zip Document.

Article and author information

Author details

  1. Jiale Zhong

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5873-0450
  2. Chen-Jun Guo

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  3. Xian Zhou

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  4. Chia-Chun Chang

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  5. Boqi Yin

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  6. Tianyi Zhang

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  7. Huanhuan Hu

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  8. Guang-Ming Lu

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    Competing interests
    The authors declare that no competing interests exist.
  9. Ji-Long Liu

    School of Life Science and Technology, ShanghaiTech University, Shanghai, China
    For correspondence
    Liujl3@shanghaitech.edu.cn
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4834-8554

Funding

Ministry of Science and Technology of the People's Republic of China (2021YFA0804701-4)

  • Ji-Long Liu

National Natural Science Foundation of China (31771490)

  • Ji-Long Liu

Shanghai Science and Technology Commission (20JC1410500)

  • Ji-Long Liu

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2022, Zhong et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,601
    views
  • 374
    downloads
  • 24
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Jiale Zhong
  2. Chen-Jun Guo
  3. Xian Zhou
  4. Chia-Chun Chang
  5. Boqi Yin
  6. Tianyi Zhang
  7. Huanhuan Hu
  8. Guang-Ming Lu
  9. Ji-Long Liu
(2022)
Structural basis of dynamic P5CS filaments
eLife 11:e76107.
https://doi.org/10.7554/eLife.76107

Share this article

https://doi.org/10.7554/eLife.76107

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Joar Esteban Pinto Torres, Mathieu Claes ... Yann G-J Sterckx
    Research Article

    African trypanosomes are the causative agents of neglected tropical diseases affecting both humans and livestock. Disease control is highly challenging due to an increasing number of drug treatment failures. African trypanosomes are extracellular, blood-borne parasites that mainly rely on glycolysis for their energy metabolism within the mammalian host. Trypanosomal glycolytic enzymes are therefore of interest for the development of trypanocidal drugs. Here, we report the serendipitous discovery of a camelid single-domain antibody (sdAb aka Nanobody) that selectively inhibits the enzymatic activity of trypanosomatid (but not host) pyruvate kinases through an allosteric mechanism. By combining enzyme kinetics, biophysics, structural biology, and transgenic parasite survival assays, we provide a proof-of-principle that the sdAb-mediated enzyme inhibition negatively impacts parasite fitness and growth.

    1. Structural Biology and Molecular Biophysics
    Manming Xu, Sarath Chandra Dantu ... Shozeb Haider
    Research Article

    The relationship between protein dynamics and function is essential for understanding biological processes and developing effective therapeutics. Functional sites within proteins are critical for activities such as substrate binding, catalysis, and structural changes. Existing computational methods for the predictions of functional residues are trained on sequence, structural, and experimental data, but they do not explicitly model the influence of evolution on protein dynamics. This overlooked contribution is essential as it is known that evolution can fine-tune protein dynamics through compensatory mutations either to improve the proteins’ performance or diversify its function while maintaining the same structural scaffold. To model this critical contribution, we introduce DyNoPy, a computational method that combines residue coevolution analysis with molecular dynamics simulations, revealing hidden correlations between functional sites. DyNoPy constructs a graph model of residue–residue interactions, identifies communities of key residue groups, and annotates critical sites based on their roles. By leveraging the concept of coevolved dynamical couplings—residue pairs with critical dynamical interactions that have been preserved during evolution—DyNoPy offers a powerful method for predicting and analysing protein evolution and dynamics. We demonstrate the effectiveness of DyNoPy on SHV-1 and PDC-3, chromosomally encoded β-lactamases linked to antibiotic resistance, highlighting its potential to inform drug design and address pressing healthcare challenges.