Yeast moclo secretion and surface display toolkit 2. 0: improvements and applications for analysis of protein–protein interactions and whole-cell biocatalysis

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dc.contributor.authorJurić, Vanja
dc.contributor.authorErwin, Leah G.
dc.contributor.authorO’Riordan, Nicola M.
dc.contributor.authorMaher, Eamonn
dc.contributor.authorHolmes, Justin D.
dc.contributor.authorYoung, Paul W.
dc.contributor.funderScience Foundation Ireland (SFI)
dc.contributor.funderIrish Research Council
dc.contributor.funderEli Lilly
dc.contributor.funderThe Biochemical Society
dc.date.accessioned2026-07-07T14:50:02Z
dc.date.available2026-07-07T14:50:02Z
dc.date.issued2026-05-15
dc.description© 2026, the Authors. Published by American Chemical Society. This publication is licensed under CC-BY 4.0.
dc.description.abstractSaccharomyces cerevisiae is an invaluable model organism for both fundamental biological research and biotechnological applications including recombinant protein production as well as protein and metabolic engineering. We previously developed a modular cloning (MoClo) based toolkit for S. cerevisiae that facilitates rapid optimization of signal peptides and anchor proteins for efficient secretion and/or surface display of heterologous proteins of interest. Here we describe further improvements and applications of this MoClo yeast secretion and display (MoClo YSD) toolkit. New parts encoding anchor proteins based on N-terminal fusion to a truncated Aga1 and C-terminal fusion to Aga2, each with three possible epitope tag options, are described. We also added parts that facilitate high throughput detection of secreted proteins of interest through GFP fluorescence complementation and parts encoding “secretion boosting” yeast proteins, whose overexpression has previously been reported to enhance secretion of heterologous proteins. In addition, two surface display applications of the toolkit are showcased. We demonstrate that yeast surface display of an anti-GFP nanobody allows cost-effective evaluation of the interactions of GFP-tagged proteins of interest, either by flow cytometry or yeast-based coimmunoprecipitation. In addition, using yeast cells as whole-cell catalysts, we show that codisplay of the poly(ethylene terephthalate) (PET) degrading enzyme leaf-branch compost cutinase with hydrophobin1 enhances the breakdown of PET plastic, while triple codisplay of these proteins with MHETase causes complete conversion of the intermediary monohydroxy-ethyl-terephthalate (MHET) to terephthalic acid. The diverse applications described herein demonstrate the broad applications of the updated MoClo YSD toolkit 2.0 in both synthetic biology and other research fields.en
dc.description.sponsorshipScience Foundation Ireland|12/RC/2278_P2) Irish Research Council|EPSPG/2020/307 Eli Lilly|Research Scholarship Biochemical Society|Summer Vacation Studentship
dc.format.extent22
dc.format.extent4097501
dc.identifier.authororcidJurić, Vanja
dc.identifier.authororcidErwin, Leah G.
dc.identifier.authororcidO’Riordan, Nicola M.
dc.identifier.authororcidMaher, Eamonn
dc.identifier.authororcidHolmes, Justin D.§0000-0001-5087-8936
dc.identifier.authororcidYoung, Paul W.§0000-0002-1307-0249
dc.identifier.citationJurić, V, Erwin, L G, O’Riordan, N M, Maher, E, Holmes, J D & Young, P W 2026, 'Yeast moclo secretion and surface display toolkit 2. 0: improvements and applications for analysis of protein–protein interactions and whole-cell biocatalysis', ACS Synthetic Biology, vol. 15, no. 5, pp. 2032-2053. https://doi.org/10.1021/acssynbio.6c00085
dc.identifier.doi10.1021/acssynbio.6c00085
dc.identifier.endpage2053
dc.identifier.issn2161-5063
dc.identifier.issued5
dc.identifier.journaltitleACS Synthetic Biology
dc.identifier.otherORCID: /0000-0001-5087-8936/work/219992614
dc.identifier.startpage2032
dc.identifier.urihttps://hdl.handle.net/10468/19015
dc.identifier.urlhttps://www.scopus.com/pages/publications/105039067680
dc.identifier.volume15
dc.language.isoeng
dc.rightscc_by
dc.subjectMoClo YSD toolkit
dc.subjectModular cloning
dc.subjectProtein interactions
dc.subjectSaccharomyces cerevisiae
dc.subjectWhole cell biocatalysis
dc.subjectYeast surface display
dc.subject[BiochemistryCellBiology]
dc.subjectBiomedical Engineering
dc.subjectBiochemistry, Genetics and Molecular Biology (miscellaneous)
dc.titleYeast moclo secretion and surface display toolkit 2. 0: improvements and applications for analysis of protein–protein interactions and whole-cell biocatalysisen
dc.typeArticle (Peer reviewed)
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