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Biotechnol."],"abstract":"<jats:title>Abstract<\/jats:title><jats:p><jats:italic>Pichia pastoris<\/jats:italic> is a popular yeast host for high-level heterologous expression of proteins on an industrial scale owing to its reliable expression, robust growth, high fermentation density, and easy genetic manipulation and cultivation at a relatively low cost. Of particular interest is its high secretion efficiency for small proteins including insulin, human serum albumin, vaccines, enzymes, and llama-derived heavy-chain only antibodies (nanobodies) for pharmaceutical and research applications. However, a recurring challenge in using <jats:italic>P. pastoris<\/jats:italic> heterologous secretory proteins is the co-purification of a sticky, yellow pigment which has been identified as a tetra-benzoyl disaccharide. Current methods for pigment removal involve crystallization of the heterologous secretory protein, active carbon absorption, and chromatography using cation exchange and hydrophobic interaction. Here, we present a simple and effective method to remove the yellow pigment, demonstrated with divalent nanobodies targeting SARS-CoV-2. The method entails capturing the nanobody on an affinity column and subsequent washing with the zwitterionic detergent lauryldimethylamine <jats:italic>N<\/jats:italic>-oxide (LDAO). We anticipate the method become generally useful to remove pigments from secretion proteins produced in <jats:italic>P. pastoris<\/jats:italic>, offering a practical solution to enhance the purity of heterologous proteins in various biotechnological applications.<\/jats:p>","DOI":"10.1007\/s44307-024-00013-z","type":"journal-article","created":{"date-parts":[[2024,2,8]],"date-time":"2024-02-08T10:02:42Z","timestamp":1707386562000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["A simple and effective method to remove pigments from heterologous secretory proteins\u00a0expressed in Pichia pastoris"],"prefix":"10.1007","volume":"2","author":[{"given":"Tingting","family":"Li","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hongmin","family":"Cai","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yanling","family":"Lai","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hebang","family":"Yao","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4729-4678","authenticated-orcid":false,"given":"Dianfan","family":"Li","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2024,2,8]]},"reference":[{"key":"13_CR1","doi-asserted-by":"publisher","first-page":"5301","DOI":"10.1007\/s00253-014-5732-5","volume":"98","author":"M Ahmad","year":"2014","unstructured":"Ahmad M, Hirz M, Pichler H, Schwab H. 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