The impact of methanol concentration on recombinant protein glycosylation in Pichia pastoris SuperMan5
Author(s)
Type
Journal Article
Abstract
The methylotrophic yeast Pichia pastoris (also known as Komagataella phaffii) is a prominent platform for recombinant protein production, offering benefits such as thermo- and osmotolerance, high-density growth, and efficient protein secretion. Its ability to metabolise methanol, an increasingly available carbon source, enhances its cost-effectiveness and sustainability for industrial use. As a eukaryotic host, P. pastoris ensures proper protein folding and post-translational modifications (PTMs), including glycosylation, which is essential for correct folding and endoplasmic reticulum (ER) quality control. While ER-transferred glycans are critical for maturation, additional modification in the Golgi apparatus can yield larger glycans whose impact on stability, solubility, and bioactivity may be either beneficial or undesirable, depending on the application of the heterologous protein. The impact of induction conditions on glycosylation of proteins secreted by P. pastoris SuperMan5 was examined, using the DS-1 (G2P[4]) and WA (G1P[8]) VP8* rotavirus capsid proteins as a model. An ELISA-based screening system was employed for clone selection and media optimization, with results showing easy integration into automated workflows. Methanol concentration was found to impact both N- and O-linked glycosylation complexity, shaping the glycosylation profile of the target protein as well as the P. pastoris secretome. This study underscores the importance of optimising cultivation conditions to enhance protein yield, refine glycosylation, and minimise impurities, all of which are crucial for large-scale production and efficient downstream processing. It also suggests a method for easy modulation of glycosylation depending on the target application and the desired level of glycosylation.
Date Issued
2025-12-01
Date Acceptance
2025-11-13
Citation
Microbial Biotechnology, 2025, 18 (12)
ISSN
1751-7907
Publisher
Wiley
Start Page
e70272
Journal / Book Title
Microbial Biotechnology
Volume
18
Issue
12
Copyright Statement
© 2025 The Author(s). Microbial Biotechnology published by John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
License URL
Identifier
https://www.ncbi.nlm.nih.gov/pubmed/41347629
Subjects
bioprocess optimization
Biotechnology & Applied Microbiology
EXPRESSION
glycosylation modulation
Life Sciences & Biomedicine
MANNOSE
Microbiology
Pichia pastoris
post-translational modifications
recombinant protein production
Science & Technology
SECRETION
STABILITY
STRATEGIES
YEAST
Publication Status
Published
Coverage Spatial
United States
Article Number
e70272
Date Publish Online
2025-12-05