Computational design and structure dynamics analysis of bifunctional chimera of endoxylanase from Clostridium thermocellum and xylosidase from Bacteroides ovatus

Development of chimeric enzymes by protein engineering can more efficiently contribute toward biomass conversion for bioenergy generation. Therefore, prior to experimental validation, a computational approach by modeling and molecular dynamic simulation can assess the structural and functional behavior of chimeric enzymes. In this study, a bifunctional chimera, Ct Xyn11A- Bo GH43A comprising an efficient endoxylanase ( Ct Xyn11A) from Clostridium thermocellum and xylosidase ( Bo GH43A) from Bacteroides ovatus was computationally designed and its binding and stability analysis with xylooligosaccharides were performed. The modeled chimera showed β-jellyroll fold for Ct Xyn11A and 5-bladed β-propeller fold for Bo GH43A module. Stereo-chemical properties analyzed by Ramachandran plot showed 98.8% residues in allowed region, validating the modeled chimera. The catalytic residues identified by multiple sequence alignment were Glu94 and Glu184 for Ct Xyn11A and Asp229 and Glu384 for Bo GH43A modules. Ct Xyn11A followed retaining-type, whereas Bo GH43A enforced inverting-type of reaction mechanism during xylan hydrolysis as revealed by superposition and GH11 and GH43 familial analyses. Molecular docking studies showed binding energy, (ΔG) − 4.54 and − 4.18 kcal/mol for Ct Xyn11A and Bo GH43A modules of chimera, respectively, with xylobiose, while − 3.94 and − 3.82 kcal/mol for Ct Xyn11A and Bo GH43A modules of chimera, respectively, with xylotriose. MD simulation of Ct Xyn11A- Bo GH43A complexed with xylobiose and xylotriose till 100 ns displayed stability by RMSD, compactness by R g and conformational stability by SASA analyses. The lowered values of RMSF in active-site residues, Glu94, Glu184, Asp229, Asp335 and Glu384 confirmed the efficient binding of chimera with xylobiose and xylotriose. These results were in agreement with the earlier experimental studies on Ct Xyn11A releasing xylooligosaccharides from xylan and Bo GH43A releasing d -xylose from xylooligosaccharides and xylobiose. The chimera showed stronger affinity in terms of total short-range interaction energy; − 190 and − 121 kJ/mol for with xylobiose and xylotriose, respectively. The bifunctional chimera, Ct Xyn11A- Bo GH43A showed stability and integrity with xylobiose and xylotriose. The designed chimera can be constructed and applied for efficient biomass conversion.