ORIGINAL RESEARCH article
Front. Microbiol.
Sec. Microbiotechnology
Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1586229
This article is part of the Research TopicSynthetic Biology for Non-Model MicrobesView all 5 articles
An Optimised Promoter and Signal Peptide Improves Methionine Production of a Genetically Engineered Candida utilis Harboring the δzein Gene
Provisionally accepted
- 1Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences, Hohhot, Inner Mongolia Autonomous Region, China
- 2Inner Mongolia Normal University, Hohhot, Inner Mongolia, China
- 3Hulunbuir Vocational Technical College, Hulunbuir, China
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L-methionine is nutritionally indispensable for humans and animals. It is widely applied to feed, livestock and poultry breeding, food, medicine, energy and chemical industries. Maize endosperm contains a stable protein called δ-zein, which is abundant in sulfur amino acids, including methionine. Candida utilis (C. utilis) has been utilized as a cell factory to express and produce recombinant products. However, there is limited information on its genetic background and expression regulatory elements. In this study, we aimed to improve methionine yields in an engineered C. utilis harboring the δ-zein gene by identifying a strong promoter and optimal signal peptide. A C. utilis glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter mutant library was constructed and screened to obtain a strong promoter. Subsequently, de novo sequencing of the C. utilis genome was performed using a combination of second-generation Illumina-Seq sequencing platform and third-generation nanopore sequencing technique. Endogenous signal peptides of C. utilis were analyzed by sequencing the C. utilis genome. Recombinant C. utilis strains with homologous integration expression vectors of different signal peptides were constructed and screened for C. utilis optimal signal peptides for secretion of δ-zein. Finally, a secretory expression system pGS-zein containing a strong promoter GP6 and an optimal signal peptide SP8 was constructed. In the food-grade engineered C. utilis C/pGS-zein methionine content increased by 21.09% compared with that of C/psP with the original promoter, and by 33.64% compared to wild-type C. utilis. This study demonstrates successful expression and secretion of δ-zein in C. utilis and establishes a foundation for enhanced methionine production of heterologous proteins in C. utilis. More importantly, these high-performance biological elements provide fundamental knowledge and technical knowhow for enhanced production of heterologous proteins in C. utilis.
Keywords: Candida utilis, Promoter, Signal peptides, Methionine, Expression vector
Received: 02 Mar 2025; Accepted: 06 Jun 2025.
Copyright: © 2025 He, Su, Ao, He, Wang, Chun and Gong. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Gaowa Gong, Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences, Hohhot, Inner Mongolia Autonomous Region, China
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