Large protein complex interfaces have evolved to promote cotranslational assembly

  1. Mihaly Badonyi
  2. Joseph A Marsh

  • Curated by eLife

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    Evaluation Summary:

    The authors use a combination of proteome-specific protein complex structures and publicly available ribosome profiling data to show that cotranslational assembly is favored by large N-terminal intermolecular interfaces. The manuscript represents an important contribution to the field of protein biosynthesis pathways by suggesting an intuitive evolutionary mechanism that can promote co-translational assembly pathways in mammalians, yeast, and bacteria.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)

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Abstract

Assembly pathways of protein complexes should be precise and efficient to minimise misfolding and unwanted interactions with other proteins in the cell. One way to achieve this efficiency is by seeding assembly pathways during translation via the cotranslational assembly of subunits. While recent evidence suggests that such cotranslational assembly is widespread, little is known about the properties of protein complexes associated with the phenomenon. Here, using a combination of proteome-specific protein complex structures and publicly available ribosome profiling data, we show that cotranslational assembly is particularly common between subunits that form large intermolecular interfaces. To test whether large interfaces have evolved to promote cotranslational assembly, as opposed to cotranslational assembly being a non-adaptive consequence of large interfaces, we compared the sizes of first and last translated interfaces of heteromeric subunits in bacterial, yeast, and human complexes. When considering all together, we observe the N-terminal interface to be larger than the C-terminal interface 54% of the time, increasing to 64% when we exclude subunits with only small interfaces, which are unlikely to cotranslationally assemble. This strongly suggests that large interfaces have evolved as a means to maximise the chance of successful cotranslational subunit binding.

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  1. Version published to 10.7554/elife.79602 on eLife
  2. eLife

    Evaluation Summary:

    The authors use a combination of proteome-specific protein complex structures and publicly available ribosome profiling data to show that cotranslational assembly is favored by large N-terminal intermolecular interfaces. The manuscript represents an important contribution to the field of protein biosynthesis pathways by suggesting an intuitive evolutionary mechanism that can promote co-translational assembly pathways in mammalians, yeast, and bacteria.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)

  3. eLife

    Reviewer #1 (Public Review):

    The authors used available protein complex structures and ribosome profiling data to analyse how the interface size between proteins in the complex and the position of the largest interface correlates with the propensity of subunits to assemble cotranslationally.

    The strength of the paper is in finding a simple well-defined parameter that may direct the evolution of protein interfaces and cotranslational protein folding. There are some weaknesses in presenting statistical significance (Fig S1C) and justification/validation of the assembly-onset mapping approach summarized in Fig. 2A. Provided that the data shown in Fig. S1C are significant, the results support the conclusions of the paper.

    The paper makes an important contribution to protein science by making a proteome-wide analysis of parameters that contribute to cotranslational folding and by finding a number of as yet unidentified candidates for simultaneous assembly, which is an important starting point for biochemical experiments to test the mechanism of folding.

  4. eLife

    Reviewer #2 (Public Review):

    The manuscript "Large protein complex interfaces have evolved to promote cotranslational assembly" by Mihaly Badonyi and Joseph A Marsh combines analysis of experimental data from ribosome profiling experiments (mainly Bertolini et al., 2021) with structural data on protein complexes to test the impact of interface size on co-translational complex assembly pathways in mammalians. The authors find a strong correlation between interface size and co-translational subunit association. Expanding their structural analysis to yeast and E.coli complexes, the authors find evidence supporting the hypothesis that large interfaces have evolved to promote co-translational assembly. Thus, larger N` terminal interfaces can serve to facilitate successful co-translational assembly interactions, protecting the nascent proteome.

    Weaknesses:

    - The correlation to yeast and bacteria experimental data regarding cotranslational assembly pathways is lacking, compared to the human analysis. As the manuscript suggests this is the basis for the hypothesis that large interfaces have evolved to promote co-translational assembly, this requires addressing.
    - Interface size differences are analyzed in different complex subsets: homomeric symmetry groups as well as hetero-oligomers. However, the statistical significance between these groups requires clarification.

    Strengths:

    - The manuscript provides a vast analysis of interface characteristics derived from structural as well as state-of-the-art modeling data utilizing AlphaFold. This analysis spans ~4000 human complexes as well as hundreds of yeast and bacteria data.
    - The manuscript furthermore provides an in-depth analysis of ribosome profiling based on experimental data in human cell lines, analyzing proteome-wide assembly interaction, initiating during protein synthesis.
    - Comparing these analyses, the authors provide novel evolutionary concepts that can be utilized to predict co-translational assembly pathways in mammalians, yeast, and bacteria.
    - The authors demonstrate how N` terminal interfaces have evolved to larger sizes, to facilitate co-translational assembly interactions.

  5. eLife

    Reviewer #3 (Public Review):

    Results presented in this manuscript demonstrate a strong correspondence between interface size and cotranslational subunit assembly mechanism. Additionally, in this manuscript, the authors have compared the sizes of first and last translated interfaces of heteromers in bacterial, yeast, and human complexes and detected a clear preference in N-terminal interfaces, which will be translated first and thus more likely to form cotranslationally, to be larger than C-terminal interfaces. The authors concluded that large interfaces have evolved as a means to maximize the chance of successful co-translational subunit assembly thus minimizing misfolding and unwanted interactions with other proteins in the cell.

    This is a very interesting study further shedding light on the mechanism and the pathway of cotranslational protein folding, which, however, prompts many additional questions as they relate to the exact nature of the cotranslationally assembling interfaces.

  6. eLife

    Reviewer #4 (Public Review):

    The authors tested several questions related to the hypothesis that larger oligomer-forming protein interfaces would promote co-translational assembly, and therefore show an evolutionary enrichment across the structural proteome of E. coli, yeast, and humans. The authors used a bioinformatic approach applied to high-throughput selective ribosome profiling data, protein structural information, as well as appropriate statistical methods in most cases.

    Regarding the conclusion that "Cotranslationally assembling subunits are characterised by large interfaces", a very serious concern is that it appears this conclusion may not be statistically valid when they apply the analysis to the 'high confidence' data. (Note well, the 'high confidence' data category was created by the lab that originally created and reported the high-throughput experimental data.) Specifically, in Fig. S1C, the authors do not provide p-values demonstrating that "both high and low confidence candidates have a larger mean area than unannotated proteins". The p-value should be reported. And if it is not significant, then it is not clear whether the data supports the conclusion.

    More generally, if the original creators of the experimental data had some data they were confident in, and some data they were less confident in, shouldn't the author's main text analyses be restricted to the 'high confidence' experimental data?

    A measure of effect size should be reported for how common it is that for a given protein, the N-terminal interface is larger than the C-terminal interface. This effect size is important so readers can get a sense of whether this is commonplace or not.

    If the conclusions hold up for the 'high confidence' data subset then this work is quite significant, as this work would then offer structural and energetic explanations for why some proteins follow a co-translational assembly route as compared to a post-translational assembly route, and how evolution has biased the distribution of these properties.

  7. Version published to 10.1101/2021.05.26.445863v2 on bioRxiv
  8. Version published to 10.1101/2021.05.26.445863v1 on bioRxiv