Computation-Aided Phylogeny-Oriented Engineering of
β‑Xylosidase: Modification of “Blades”
to Enhance Stability and Activity for the Bioconversion of Hemicellulose
to Produce Xylose
posted on 2024-01-26, 05:29authored byChenchen Zhang, Wenjing Gao, Zhaolin Song, Mengjun Dong, Huixin Lin, Gang Zhu, Mengka Lian, Yunjie Xiao, Fuping Lu, Fenghua Wang, Yihan Liu
Hemicellulose
is a highly abundant, ubiquitous, and renewable natural
polysaccharide, widely present in agricultural and forestry residues.
The enzymatic hydrolysis of hemicellulose has generally been accomplished
using β-xylosidases, but concomitantly increasing the stability
and activity of these enzymes remains challenging. Here, we rationally
engineered a β-xylosidase from Bacillus clausii to enhance its stability by computation-aided design combining ancestral
sequence reconstruction and structural analysis. The resulting combinatorial
mutant rXYLOM25I/S51L/S79E exhibited highly improved robustness,
with a 6.9-fold increase of the half-life at 60 °C, while also
exhibiting improved pH stability, catalytic efficiency, and hydrolytic
activity. Structural analysis demonstrated that additional interactions
among the propeller blades in the catalytic module resulted in a much
more compact protein structure and induced the rearrangement of the
opposing catalytic pocket to mediate the observed improvement of activity.
Our work provides a robust biocatalyst for the hydrolysis of agricultural
waste to produce various high-value-added chemicals and biofuels.