Improving mannanase production in Bacillus subtilis for fibre hydrolysis during solid-state fermentation of palm kernel meal

• Published in: Biochemical engineering journal Vol. 212; p. 109479
• Main Authors: Ong, Wei Li, Li, Zhi, Ng, Kian-Hong, Zhou, Kang
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 01-12-2024
• Subjects: Bacillus subtilis; Mannanase; Palm kernel meal; Solid-state fermentation; Palm kernel meal; Solid-state fermentation; Bacillus subtilis; Mannanase
• ISSN: 1369-703X
• DOI: 10.1016/j.bej.2024.109479

The primary challenge in utilizing palm kernel meal (PKM, an agricultural by-product) as non-ruminant livestock feed is its high fibre content, predominantly in the form of mannan. Microbial fermentation offers an economically favourable alternative to enzyme supplementation for breaking down fibre in lignocellulosic biomass. In a recent study, our group isolated a B. subtilis strain F6 with a fast response time for mannanase production upon exposure to PKM. This work focuses on improving the mannanase production of the B. subtilis strain to achieve greater fibre hydrolysis of PKM without extending fermentation time. Mannanase GmuG, sourced from B. subtilis F6 and verified for its hydrolytic activity on PKM fibre, was homologously expressed using a replicative plasmid (pBE-S). Enzyme production was systematically improved by optimizing various regulatory elements, including the promoter, ribosome binding site, and signal peptide. Consequently, the neutral detergent fibre content of PKM was substantially reduced by 36.4 % in 22 h of solid-state fermentation using the engineered strain. Lastly, the highest mannanase-producing strain was examined for scaled-up fermentation. The impacts of fermentation on fibre and protein contents, as well as the surface morphology of PKM, were analysed. The outcomes of this study offer an efficient method for robust mannanase expression in B. subtilis and its potential application in the biotransformation of PKM and other mannan-rich bioresources for improved feed utilization.
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•Effective fibre hydrolysis relies on strain selection and robust enzyme expression.•Strain variations are crucial for adapting to unconventional substrates.•Over 60-fold mannanase enhancement via plasmid optimization in B. subtilis.•Mannanase expression improved fibre hydrolysis from 10 % to 36 %.•Transcriptome data aids promoter identification under specific growth conditions.