An iris-like pore for protein import into peroxisomes

D. He; Y. Gao; T. Rapoport

doi:10.64898/2026.10.23.761396

AI summary · DOI: 10.64898/2026.10.23.761396 · bioRxiv: Cell Biology

An iris-like pore for protein import into peroxisomes

D. He, Y. Gao, T. Rapoport

Original paper Published on 27 Oct. 2026 bioRxiv preprint License: CC BY 4.0 Also in: fr

How to read this summary? Illustrated guide

The essentials

The authors propose that a 12-subunit PEX13 pore changes diameter through iris-like helix sliding, potentially enabling peroxisomes to import folded cargo of different sizes.

Question

How is the peroxisomal import conduit organized, and how can it accommodate folded proteins and oligomers of different sizes? The study asks whether PEX13 forms a pore that can change its diameter.

Method

The authors expressed human PEX13 in Pichia pastoris, purified it, and analyzed wild-type and mutant assemblies by cryo-EM; yeast PEX13 was also tested in import assays and disulfide-crosslinking experiments. Cryo-EM resolved wild-type pores of approximately 52 Å and 70 Å at about 6.2 Å overall resolution, while a three-acidic-residue mutant yielded a 2.8 Å map of an approximately 52 Å pore; an approximately 100 Å pore was predicted by AlphaFold and assessed with crosslinking.

Findings

Result PEX13 assembles into a pore ring of 12 molecules, with alternating amphipathic helices forming the wall around the YG phase.
See the passage in the paper
The maps show a ring of twelve left-tilted helices surrounding an almost circular pore.
Result The wild-type PEX13 structures show pores of approximately 52 Å and 70 Å, while the larger approximately 100 Å pore is predicted rather than experimentally structurally resolved.
See the passage in the paper
In two maps, the pore diameter is ∼70 Å, and in the third ∼52 Å.
Result Crosslinking results support the proposal that neighboring helices slide relative to one another to produce different pore configurations.
See the passage in the paper
Together, these data confirm the different channels generated by yeast and human PEX13.
Authors’ interpretation The authors propose that the pore can widen in an iris-like motion, potentially allowing folded and oligomeric cargo to pass.
See the passage in the paper
By contrast, our results suggest that the peroxisomal import channel has a unique cargo-adaptable pore that could allow folded and even oligomeric cargo to pass through 51.
Result Mutating three conserved acidic residues in the amphipathic helix produces an import-inactive channel with a pore resembling the smaller wild-type state.
See the passage in the paper
The tilt angle of the AHs (52°) and the pore diameter (∼52 Å) are similar to those of the small wild-type channel (53° and ∼52 Å, respectively).

Limitations

The proposed transition between pore states was not directly visualized, and the approximately 100 Å pore remains a prediction.
See the passage in the paper
Although we have not directly visualized the transitions between the different channel states, they likely occur by a simple iris-like motion of the AHs.
It remains to be tested experimentally whether receptor and large-cargo passage induces channel widening.
See the passage in the paper
Although challenging, it will be important to test experimentally whether channel widening is induced when PEX5 and large cargo move through.
The authors do not establish whether pore sizes are limited to discrete states or include intermediate configurations.
See the passage in the paper
Whether the channel adopts only discrete pore sizes (∼52 Å, ∼70 Å, ∼100 Å) is unclear.

Why it matters

The authors argue that PEX13 provides a cargo-adaptable import pore, a proposed translocation mechanism distinct from conventional aqueous channels and locally thinned membranes. They suggest that its resemblance to the nuclear pore may point to shared solutions for transporting folded proteins between compartments.

Written by AI from the paper. Passages are checked word for word against the text we analysed; the rest may contain errors.

AI-generated keywords: Peroxisomes PEX13 protein import cryo-EM membrane channels

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Comprehensive Summary
Layman's Summary
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Original abstract

The paper investigates how peroxisomes import folded proteins, including cargoes that may exceed the apparent size of the known import machinery’s passageway. Peroxisomal matrix proteins bind soluble receptors such as PEX5, which dock at the membrane and pass through a conduit involving PEX13. Previous work implicated PEX13’s tyrosine/glycine-rich (YG) region in forming a selective phase, but the channel’s membrane architecture and capacity to accommodate cargoes of different sizes were unknown. The authors purified human PEX13 from *Pichia pastoris* and used single-particle cryo-electron microscopy (cryo-EM) to determine structures of its oligomeric assemblies. They also used cysteine-based disulfide crosslinking in human and yeast PEX13 to test contacts between helices and assess their orientations. Finally, they tested the effects of mutations on protein import in yeast. Wild-type cryo-EM reconstructions, with an overall resolution of about 6.2 Å, revealed rings of 12 PEX13 molecules forming channels with pore diameters of approximately 52 Å or 70 Å. The channel walls consist of tilted amphipathic helices, with their hydrophobic faces toward the surrounding membrane and their hydrophilic faces toward the pore. The helices alternate in membrane orientation, consistent with PEX13’s dual topology. Experiments also showed that disrupting the YG region impairs oligomerization, supporting a role for the YG phase in stabilizing the channel. A higher-resolution structure of an import-inactive human PEX13 mutant—in which three conserved acidic residues were replaced with alanines—revealed a 12-helix channel with a pore approximately 52 Å across. The reconstruction reached 2.8 Å resolution with threefold symmetry imposed and 3.3 Å without it. In yeast, the corresponding triple mutant was essentially inactive in the import assay. Mutating one of the three acidic residues had no detectable effect, while mutating two reduced import. The mutant structure resembles the smaller wild-type pore, and the altered charges point into the pore. The authors propose that electrostatic repulsion between the acidic residues helps widen the channel, although this mechanism was not directly demonstrated. The authors also propose a channel approximately 100 Å in diameter, based on AlphaFold2 predictions rather than an experimentally determined structure. In their model, pairs of neighboring helices form relatively fixed units, while small-residue-rich interfaces allow adjacent units to slide. This iris-like movement changes helix tilt and pore size. Crosslinking at selected positions in yeast and human PEX13 supported contacts predicted for the 52 Å, 70 Å, and 100 Å states. Mutations that introduced aromatic residues at the fixed interface or a bulky residue at the variable interface abolished yeast import, consistent with these interfaces being functionally important. The authors propose that the 52 Å pore represents a resting state. The 70 Å pore could accommodate PEX5’s cargo-binding TPR domain and some cargoes, while a pore close to 100 Å could allow large oligomers to pass. Overall, the findings support a model in which PEX13 forms a membrane-embedded, size-adjustable passageway for folded and potentially oligomeric cargo, rather than relying solely on a narrow aqueous pore or local membrane thinning. Important limitations remain. The 100 Å state was predicted rather than structurally observed, and transitions between pore states were not directly visualized. The proposed roles of cargo, receptor binding, and acidic-residue repulsion in triggering channel expansion remain hypotheses. In addition, the comparatively low resolution of the wild-type structures limits atomic-level interpretation.

An Iris-Like Pore for Protein Import into Peroxisomes

Peroxisomes are cellular compartments that carry out important chemical reactions, and they face a remarkable logistical challenge: many of the proteins they need arrive already folded. How can a large, structured protein cross the peroxisome’s membrane? In “An iris-like pore for protein import into peroxisomes,” D. He, Y. Gao, and T. Rapoport investigate a proposed solution: a membrane channel made from PEX13 that may widen to accommodate cargo of different sizes.

A passageway with a flexible design

Peroxisomal matrix proteins bind soluble receptors, including PEX5. The receptor and its cargo then dock at the membrane, where PEX13 is part of the import machinery. Earlier work suggested that a tyrosine/glycine-rich, or YG, region of PEX13 helps form a selective phase involved in import. But the channel’s membrane architecture—and how it could handle cargoes larger than a narrow pore—remained unclear.

To address this, the researchers purified human PEX13 and used cryo-electron microscopy to examine its assemblies. They also used cysteine-based disulfide crosslinking in human and yeast PEX13 to test whether selected parts of the protein came into contact. Finally, they tested how mutations affected protein import in yeast.

Ring-shaped channels and an adaptable pore

The wild-type cryo-EM reconstructions revealed rings containing 12 PEX13 molecules. These rings formed channels with pore diameters of about 52 or 70 angstroms. Their walls are made of tilted amphipathic helices: hydrophobic surfaces face the surrounding membrane, while hydrophilic surfaces face the channel’s interior. The helices alternate in membrane orientation, consistent with PEX13’s dual topology.

The YG region also appears to matter structurally. Disrupting it impaired PEX13 oligomerization, supporting the idea that the YG phase helps stabilize the assembled channel.

For a more detailed view, the team examined a human PEX13 mutant in which three conserved acidic residues were changed to alanines. This mutant was inactive in import tests, and its structure revealed a 12-helix channel with a pore approximately 52 angstroms wide. The reconstruction reached 2.8-angstrom resolution with threefold symmetry imposed, and 3.3 angstroms without it. In yeast, the corresponding triple mutant was also essentially inactive. Changing just one of the acidic residues had no detectable effect, while changing two reduced import.

The acidic residues point toward the pore. The researchers suggest that repulsion between their negative charges could help push the channel into a wider configuration. However, this explanation remains a hypothesis: the experiments did not directly demonstrate that electrostatic repulsion drives pore expansion.

How might the channel widen?

The study’s proposed largest pore, about 100 angstroms across, was not observed in a cryo-EM structure. Instead, it comes from an AlphaFold2-based model. In this model, pairs of neighboring helices form relatively fixed units, while interfaces rich in small residues let adjacent units slide past one another. The resulting motion resembles an iris opening: changes in helix tilt could enlarge or shrink the pore.

Crosslinking experiments in yeast and human PEX13 supported contacts predicted for the 52-, 70-, and 100-angstrom arrangements. Mutations that placed aromatic residues at the proposed fixed interface, or a bulky residue at the variable interface, abolished import in yeast. These results are consistent with the interfaces being important for function, although they do not directly show the channel moving between states.

What the pore sizes could mean

The researchers propose that the 52-angstrom pore is a resting state. A 70-angstrom opening could fit PEX5’s cargo-binding TPR domain and some protein cargoes, while a pore approaching 100 angstroms might allow larger oligomers to pass. If correct, this would offer a way for peroxisomes to import folded—and potentially assembled—proteins without depending only on a narrow aqueous tunnel or local thinning of the membrane.

Wild-type PEX13 forms 12-subunit rings with observed pores of about 52 and 70 angstroms.
The YG region contributes to oligomer formation, and mutations at proposed helix interfaces disrupt import.
A pore near 100 angstroms is a model-based proposal, not an experimentally resolved structure.

What remains unknown

The results support a size-adjustable PEX13 passageway, but important questions remain. The 100-angstrom state has not been structurally observed, and transitions between pore sizes have not been visualized. The proposed roles of cargo, PEX5 binding, and acidic-residue repulsion in triggering expansion still need to be tested. In addition, the relatively low resolution of the wild-type structures limits detailed interpretation of their molecular arrangements.

Even with these caveats, the study presents an intriguing picture of protein import: rather than serving as a fixed-size opening, PEX13 may form a dynamic, iris-like channel. That model offers a possible explanation for how peroxisomes can receive folded proteins of varied sizes—and a framework for future experiments to determine exactly when and how the pore opens.

Created on 02 Nov. 2026

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Weekly letter Presented in "Biology and medicine this week" of 2 November 2026. Biology and medicine this week: Hidden partners, adjustable gates and overlooked disease

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An iris-like pore for protein import into peroxisomes

The essentials

Question

Method

Findings

Limitations

Why it matters

Read further

Definitions

An Iris-Like Pore for Protein Import into Peroxisomes

A passageway with a flexible design

Ring-shaped channels and an adaptable pore

How might the channel widen?

What the pore sizes could mean

What remains unknown

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