{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T23:48:53Z","timestamp":1792108133881,"version":"4.4.0"},"reference-count":41,"publisher":"Wiley","issue":"8","license":[{"start":{"date-parts":[[2026,8,23]],"date-time":"2026-08-23T00:00:00Z","timestamp":1787443200000},"content-version":"vor","delay-in-days":22,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2026,8,1]],"date-time":"2026-08-01T00:00:00Z","timestamp":1785542400000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32401209"],"award-info":[{"award-number":["32401209"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biotechnology Journal"],"published-print":{"date-parts":[[2026,8]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Methanol is a promising renewable C1 feedstock for sustainable single\u2010cell protein (SCP) production. However, its inherent cytotoxicity and metabolic trade\u2010offs between cell growth and protein synthesis remain significant bottlenecks. Here, we established an \u201cevolutionary\u2010rational\u201d dual\u2010driven paradigm to construct a high\u2010yield\n                    <jats:italic>Pichia pastoris<\/jats:italic>\n                    chassis. Through UV mutagenesis and adaptive laboratory evolution, we developed a highly tolerant strain A40, capable of growing in 70\u00a0g\/L methanol. Notably, at 30\u00a0g\/L methanol, A40 achieved a 3.4\u2010fold higher maximum biomass than the wild\u2010type. Whole\u2010genome resequencing and reverse genetics revealed that this superior performance stems from a multi\u2010gene synergistic network rather than a single dominant mutation. To further optimize SCP production, we rationally co\u2010overexpressed nitrogen assimilation genes (\n                    <jats:italic>GLN1<\/jats:italic>\n                    ,\n                    <jats:italic>GDH1<\/jats:italic>\n                    ) and a translation elongation factor (\n                    <jats:italic>PpeEF3<\/jats:italic>\n                    ) in the A40 background. This targeted metabolic engineering effectively redirected carbon flux toward protein biosynthesis. The engineered strain A40\u20102Ge3 achieved a peak intracellular crude protein content of 67.9% and a 51.3\u00a0g\/L total titer in a 5\u2010L bioreactor, representing a 23.1% increase over the wild\u2010type strain. Collectively, this study provides deep insights into the synergistic mechanisms of methanol adaptation and establishes an efficient, scalable strategy for sustainable SCP production from C1 feedstocks.\n                  <\/jats:p>","DOI":"10.1002\/biot.70300","type":"journal-article","created":{"date-parts":[[2026,8,24]],"date-time":"2026-08-24T06:46:09Z","timestamp":1787553969000},"source":"Crossref","is-referenced-by-count":1,"title":["Evolutionary Adaptation and Targeted Metabolic Engineering Synergistically Improve Methanol\u2010Derived Single\u2010Cell Protein Production in\n                    <i>Pichia pastoris<\/i>"],"prefix":"10.1002","volume":"21","author":[{"given":"Chong","family":"Xie","sequence":"first","affiliation":[{"name":"Center For Future Food Muyuan Laboratory  Zhengzhou China"},{"name":"School of Chemical Engineering Zhengzhou University  Zhengzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chengchao","family":"Zhu","sequence":"additional","affiliation":[{"name":"Center For Future Food Muyuan Laboratory  Zhengzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junze","family":"Liu","sequence":"additional","affiliation":[{"name":"Center For Future Food Muyuan Laboratory  Zhengzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiqi","family":"Sun","sequence":"additional","affiliation":[{"name":"School of Engineering Westlake Center of Synthetic Biology and Integrated Bioengineering Westlake University  Hangzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jingliang","family":"Xu","sequence":"additional","affiliation":[{"name":"Center For Future Food Muyuan Laboratory  Zhengzhou China"},{"name":"School of Chemical Engineering Zhengzhou University  Zhengzhou 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