Engineered Folding and Secretion for Expression Improvement of α‐Amylase in Komagataella Phaffii

• DOI: 10.1002/fbe2.70005
• Publication Date: 2025-03-23T17:26:19Z
• Authors: Yilun Zhang; Xingbin Wang; Bei Han; Chaoying Yao; Qi Liu; Menghao Cai
• Keywords: Folding (DSP implementation); Expression (computer science)

ABSTRACT α‐Amylase is the second most widely produced enzyme globally, with diverse applications in fields of food, pharmaceutical, bioenergy, papermaking, etc. However, natural α‐amylase often fails to withstand extreme conditions encountered industrial processes, such as low pH and high temperatures. Previous studies identified an derived from deep‐sea sources resistance pH, subsequent amino acid mutations well enhanced its thermal stability. Nevertheless, advantageous mutant exhibited expression levels Escherichia coli , highlighting need for a more suitable host. In this study, engineered host, Komagataella phaffii was involved heterologous production α‐amylase. High‐efficiency signal peptides were screened multi‐copy integrant strains constructed achieve high‐yield strain. A total 31 key chaperones 11 vesicle transport factors further investigated facilitate protein folding secretion, which resulted 3.4‐fold increase production. Finally, batch fermentation 3‐L bioreactor achieved maximum activity 2.5 × 10 4 U/mL. This study demonstrates development strain potential applications, offering valuable insights strategies engineering high‐producing other enzymes.