Improving the catalytic performance of xylanase from Bacillus circulans through structure-based rational design

Simultaneously Enhanced Thermostability and Catalytic Activity of Xylanase from Streptomyces rameus L2001 by Rigidifying Flexible Regions in Loop Regions of the N-Terminus

“…The challenge was to seek a balance between the rigidity and flexibility of the protein structure so as to simultaneously improve the thermal stability and catalytic activity of the enzyme. , Enhancing the thermal stability of xylanases requires a more rigid structure, whereas maintaining or even increasing their catalytic activity requires a flexible structure . The GH11 family of xylanases has more flexible amino acid residues in the α-helix as well as the N-terminus, which are sensitive to thermal-induced structure unfolding.…”

“…The flexible regions of xylanase are located in the N-terminal loop region, palm area, finger area, and α-helix . The flexible sites in the structure of mesophilic xylanase can be predicted by computer-aided methods, such as B-factor analysis and molecular dynamics (MD) simulation. , Construction of extra interactions in the flexible region predicted by MD simulations is a practical strategy to improve the structural rigidity as well as the thermal stability of mesophilic xylanases. − Using sequence alignment to locate conserved sequences of thermophilic xylanases can provide guidance for the rational design of flexible regions in mesophilic xylanases. , It is postulated that conserved sequences detected by sequence alignment carry information on common features of enzymes derived from the same family . Mining conserved sequences of thermophilic xylanases can provide a mutational library for the modification of mesophilic xylanases. , Rational design of xylanase modification strategies, based on a mutational library constructed from conserved sequences located by sequence alignment in the flexible region predicted by MD simulations, is a promising approach to enhancing the thermal stability of an enzyme.…”

“…11−13 Using sequence alignment to locate conserved sequences of thermophilic xylanases can provide guidance for the rational design of flexible regions in mesophilic xylanases. 8,14 It is postulated that conserved sequences detected by sequence alignment carry information on common features of enzymes derived from the same family. 15 Mining conserved sequences of thermophilic xylanases can provide a mutational library for the modification of mesophilic xylanases.…”

Engineering of xylanases for the development of biotechnologically important characteristics

“…Here, the selection of residues in hyperthermophilic GH10 family xylanase XynAF1 has been performed based on B‐factor values determined by B‐Fitter software to improve its thermostability by site‐saturation mutagenesis (Li et al, 2021). Similarly, using flexibility analysis by HingeProt and FIRST servers to estimate the residues for substitutions, the catalytic performance of Bacillus circulans GH11 xylanase has been improved via site‐saturation mutagenesis (Min et al, 2021).…”

“…Some computational methods (e.g., B‐Fitter, Hotspot Wizard34) are used to increase the thermostability of xylanases, especially through rational‐based approaches for the selection of the flexible region and amino acids of the enzymes. Recently, the integration of the structure‐based methodology with the sequence‐based methodology has allowed the emergence of a structure‐guided consensus strategy for the determination of the target amino acid position(s) of proteins for site‐directed mutagenesis (Li et al, 2021; Min et al, 2021; Xing et al, 2021). On the other hand, since directed evolution does not require knowledge of structure and mechanism, the use of computational methods is only used to confirm the effect of mutations underlying desired characters.…”