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It is well established that folded, cofactor-containing, and even oligomeric matrix proteins can pass the peroxisomal membrane. Here, we studied whether relatively long unfolded proteins can be imported into peroxisomes using the yeast\n                    <jats:italic>Hansenula polymorpha<\/jats:italic>\n                    as a model organism. First, we designed fusion proteins containing parts of the intrinsically disordered N-terminal region of 630 residues of\n                    <jats:italic>Saccharomyces cerevisiae<\/jats:italic>\n                    nucleoporin 1 (Nsp1). These included fusion proteins containing the first N-terminal 301 or 601 residues or two times the 601 residues region of Nsp1. Green fluorescent protein was added at the N-terminus, and a peroxisomal targeting signal 1 at the C-terminus. Fluorescence microscopy revealed that all three fusion proteins colocalized with a peroxisomal marker protein, indicating that large unfolded protein domains can be imported into peroxisomes. To obtain a fully unfolded protein, we replaced the folded GFP tag by a double human influenza hemagglutinin tag, which unlike GFP is unable to fold. An in vivo protease protection assay showed that a portion of the produced proteins localized to peroxisomes, which was confirmed by quantitative immuno-electron microscopy analysis. On the basis of our observations, we conclude that a fully unfolded protein of over 1200 residues (almost 500\u00a0nm in length) can pass the peroxisomal membrane.\n                  <\/jats:p>","DOI":"10.1007\/s00418-026-02483-9","type":"journal-article","created":{"date-parts":[[2026,5,12]],"date-time":"2026-05-12T11:14:35Z","timestamp":1778584475000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["The peroxisomal importomer can accommodate an intrinsically disordered protein of 1247 residues"],"prefix":"10.1007","volume":"164","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7924-5814","authenticated-orcid":false,"given":"Marc P.","family":"Pedersen","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Arjen M.","family":"Krikken","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rinse","family":"de Boer","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7165-9679","authenticated-orcid":false,"given":"Ida J.","family":"van der Klei","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,5,12]]},"reference":[{"key":"2483_CR1","doi-asserted-by":"publisher","first-page":"225","DOI":"10.1529\/biophysj.106.091561","volume":"92","author":"SRK Ainavarapu","year":"2007","unstructured":"Ainavarapu SRK, Bruji\u0107 J, Huang HH et al (2007) Contour length and refolding rate of a small protein controlled by engineered disulfide bonds. 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