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However, the applicability of this system, particularly in food and pharmaceutical products, is limited by methanol's toxic and pro\u2010inflammatory properties. Therefore, obtaining a novel methanol\u2010free expression system is necessary. In this study, we obtained a novel expression plasmid, pHSP\u03b1A, carrying the\n                    <jats:italic>HSP70<\/jats:italic>\n                    promoter (\n                    <jats:italic>P<\/jats:italic>\n                    <jats:sub>\n                      <jats:italic>HSP70<\/jats:italic>\n                    <\/jats:sub>\n                    ) to regulate heterologous expression through heat induction. The extracellular expression of azurin was achieved using this methanol\u2010free system under the control of\n                    <jats:italic>P<\/jats:italic>\n                    <jats:sub>\n                      <jats:italic>HSP70<\/jats:italic>\n                    <\/jats:sub>\n                    , induced by either magnetic or non\u2010magnetic heating. To enhance heat\u2010induced expression, recombinant cells were immobilised with Fe\n                    <jats:sub>3<\/jats:sub>\n                    O\n                    <jats:sub>4<\/jats:sub>\n                    @PEI\n                    <jats:sub>25 kDa<\/jats:sub>\n                    nanoparticles, which facilitated heat release under an AC magnetic field, thereby increasing cell permeability and protein secretion. A time\u2010dependent increase in protein expression was observed in non\u2010magnetic heating but not under magnetic heating. However, immobilised cells exhibited a higher protein secretion capacity compared to non\u2010immobilised cells. These findings suggest that the novel methanol\u2010free expression system represents a promising alternative for heterologous gene expression, particularly for the production of therapeutically relevant and food\u2010grade recombinant proteins.\n                  <\/jats:p>","DOI":"10.1111\/1751-7915.70183","type":"journal-article","created":{"date-parts":[[2025,6,14]],"date-time":"2025-06-14T08:20:37Z","timestamp":1749889237000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Methanol\u2010Free Protein Expression in\n                    <i>Komagataella phaffii<\/i>\n                    With Magnetic or Non\u2010Magnetic Heating"],"prefix":"10.1111","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6354-2654","authenticated-orcid":false,"given":"Ibrahim","family":"Dagci","sequence":"first","affiliation":[{"name":"Department of Molecular Biology and Genetics Institute of Science and Technology, Atat\u00fcrk University  Erzurum T\u00fcrkiye"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Seyda Yildiz","family":"Arslan","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics Institute of Science and Technology, Atat\u00fcrk University  Erzurum T\u00fcrkiye"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kubra","family":"Solak","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics Faculty of Science, Atat\u00fcrk University  Erzurum T\u00fcrkiye"},{"name":"East Anatolia High Technology Application and Research Center (DAYTAM) Atat\u00fcrk University  Erzurum T\u00fcrkiye"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Melek","family":"Acar","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics Institute of Science and Technology, Atat\u00fcrk University  Erzurum 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