Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2025 Apr;197(4):2579-2597.
doi: 10.1007/s12010-024-05145-5. Epub 2025 Jan 8.

Effect of Fold-Promoting Mutation and Signal Peptide Screening on Recombinant Glucan 1,4-Alpha-maltohydrolase Secretion in Pichia pastoris

Siyi Wang  1 , Kai Zhu  1 , Pulin Liu  2
Affiliations

Effect of Fold-Promoting Mutation and Signal Peptide Screening on Recombinant Glucan 1,4-Alpha-maltohydrolase Secretion in Pichia pastoris

Siyi Wang et al. Appl Biochem Biotechnol. 2025 Apr.

Abstract

Glucan 1,4-alpha-maltohydrolase (3.2.1.133, GMH) is an important biocatalyst in the baking industry, which could delay the retrogradation of bread and improve its cold-storage durability. In the present study, a newly cloned Thgmh was characterized and secreted by Pichia pastoris (Komagataella pastoris). After computationally assisted rational design that promotes peptide folding, the maltogenic activity in supernatant was enhanced 1.6-fold in comparison with the base strain. The signal leading sequence screening and the gene dosage increment further improved secretion by approximately 6.4-fold. The purified rationally designed ThGMHs exhibited maximal activity against soluble starch at pH 7.0 and 60 ℃, and maltose is the main catalytic product. In a 5-L bioreactor, conventional fed-batch fermentation resulted in 6130 U mL-1 extracellular maltogenic activity. Therefore, a promising strain for GMH production was developed, which provides a useful reference for the secretory production of other industrial enzymes.

Keywords: Pichia pastoris; Fermentation; Glucan 1,4-alpha-maltohydrolase; Rational design; Signal leading sequence.

PubMed Disclaimer

Conflict of interest statement

Declarations. Ethics Approval: This study does not contain any experiments involving human or animal subjects. Consent to Participate: Informed consent was obtained from all individual participants included in this study. Consent to Publish: The authors agreed to publish this paper in Applied Biochemistry and Biotechnology. Competing Interests: The authors declare no competing interests.

References

    1. Miao, M., & BeMiller, J. N. (2023). Enzymatic approaches for structuring starch to improve functionality. Annual Review of Food Science and Technology, 14, 271–295. https://doi.org/10.1146/annurev-food-072122-023510 - DOI - PubMed
    1. Srichuwong, S., & Jane, J. L. (2007). Physicochemical properties of starch affected by molecular composition and structures: A review. Food Science and Biotechnology, 16, 663–674.
    1. Miao, M., Jiang, B., Jin, Z. Y., & BeMiller, J. N. (2018). Microbial starch-converting enzymes: Recent insights and perspectives. Comprehensive Reviews in Food Science and Food Safety, 17, 1238–1260. https://doi.org/10.1111/1541-4337.12381 - DOI - PubMed
    1. BeMiller, J. N., & Huber, K. C. (2015). Physical modification of food starch functionalities. Annual Review of Food Science and Technology, 6, 19–69. https://doi.org/10.1146/annurev-food-022814-015552 - DOI - PubMed
    1. Bangar, S. P., Ashogbon, A. O., Singh, A., Chaudhary, V., & Whiteside, W. S. (2022). Enzymatic modification of starch: A green approach for starch applications. Carbohydrate Polymers, 287, 119265. https://doi.org/10.1016/j.carbpol.2022.119265 - DOI

MeSH terms

Substances

Supplementary concepts

LinkOut - more resources