Abstract

Increasing numbers of small proteins with diverse physiological roles are being identified and characterized in both prokaryotic and eukaryotic systems, but the origins and evolution of these proteins remain unclear. Recent genomic sequence analyses in several organisms suggest that new functions encoded by small open reading frames (sORFs) may emerge de novo from noncoding sequences. However, experimental data demonstrating if and how randomly generated sORFs can confer beneficial effects to cells are limited. Here we show that by up-regulating hisB expression, de novo small proteins (≤ 50 amino acids in length) selected from random sequence libraries can rescue Escherichia coli cells that lack the conditionally essential SerB enzyme. The recovered small proteins are hydrophobic and confer their rescue effect by binding to the 5' end regulatory region of the his operon mRNA, suggesting that protein binding promotes structural rearrangements of the RNA that allow increased hisB expression. This study adds RNA regulatory elements as another interacting partner for de novo proteins isolated from random sequence libraries, and provides further experimental evidence that small proteins with selective benefits can originate from the expression of nonfunctional sequences.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting file. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository (Perez-Riverol et al. 2019) with the dataset identifier PXD014049.

The following data sets were generated

Article and author information

Author details

  1. Arianne M Babina

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4635-8396
  2. Serhiy Surkov

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  3. Weihua Ye

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  4. Jon Jerlström-Hultqvist

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  5. Mårten Larsson

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  6. Erik Holmqvist

    Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  7. Per Jemth

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1516-7228
  8. Dan I Andersson

    Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden
    For correspondence
    Dan.Andersson@imbim.uu.se
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6640-2174
  9. Michael Knopp

    Genome Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany
    For correspondence
    Knopp@embl.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8218-3263

Funding

Knut och Alice Wallenbergs Stiftelse (2015.0069)

  • Dan I Andersson

Vetenskapsrådet (2017-01527)

  • Dan I Andersson

Vetenskapsrådet (2019-00666)

  • Michael Knopp

Vetenskapsrådet (2020‐04395)

  • Per Jemth

Knut och Alice Wallenbergs Stiftelse (2017.0071)

  • Mårten Larsson

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

Reviewing Editor

  1. Joseph T Wade, New York State Department of Health, United States

Version history

  1. Received: March 1, 2022
  2. Preprint posted: April 15, 2022 (view preprint)
  3. Accepted: March 14, 2023
  4. Accepted Manuscript published: March 15, 2023 (version 1)
  5. Version of Record published: March 31, 2023 (version 2)

Copyright

© 2023, Babina 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.

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  1. Arianne M Babina
  2. Serhiy Surkov
  3. Weihua Ye
  4. Jon Jerlström-Hultqvist
  5. Mårten Larsson
  6. Erik Holmqvist
  7. Per Jemth
  8. Dan I Andersson
  9. Michael Knopp
(2023)
Rescue of Escherichia coli auxotrophy by de novo small proteins
eLife 12:e78299.
https://doi.org/10.7554/eLife.78299

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https://doi.org/10.7554/eLife.78299