title: Molecular modeling and MM-PBSA free energy analysis of endo-1,4-β-xylanase from Ruminococcus albus 8.
abstract: Endo-1,4-β-xylanase (EC 3.2.1.8) is the enzyme from Ruminococcus albus 8 (R. albus 8) (Xyn10A), and catalyzes the degradation of arabinoxylan, which is a major cell wall non-starch polysaccharide of cereals. The crystallographic structure of Xyn10A is still unknown. For this reason, we report a computer-assisted homology study conducted to build its three-dimensional structure based on the known sequence of amino acids of this enzyme. In this study, the best similarity was found with the Clostridium thermocellum (C. thermocellum) N-terminal endo-1,4-β-D-xylanase 10 b. Following the 100 ns molecular dynamics (MD) simulation, a reliable model was obtained for further studies. Molecular Mechanics/Poisson-Boltzmann Surface Area (MM-PBSA) methods were used for the substrate xylotetraose having the reactive sugar, which was bound in the -1 subsite of Xyn10A in the 4C1 (chair) and 2SO (skew boat) ground state conformations. According to the simulations and free energy analysis, Xyn10A binds the substrate with the -1 sugar in the 2SO conformation 39.27 kcal·mol(-1) tighter than the substrate with the sugar in the 4C1 conformation. According to the Xyn10A-2SO Xylotetraose (X4(sb) interaction energies, the most important subsite for the substrate binding is subsite -1. The results of this study indicate that the substrate is bound in a skew boat conformation with Xyn10A and the -1 sugar subsite proceeds from the 4C1 conformation through 2SO to the transition state. MM-PBSA free energy analysis indicates that Asn187 and Trp344 in subsite -1 may an important residue for substrate binding. Our findings provide fundamental knowledge that may contribute to further enhancement of enzyme performance through molecular engineering.
authors.affiliation: Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, Jilin University, Changchun 130023, China. zdlgale@126.com.
authors.forename: Dongling
authors.initials: D
authors.lastname: Zhan
authors.affiliation: Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, Jilin University, Changchun 130023, China. gykfamily@163.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Yu
authors.affiliation: Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, Jilin University, Changchun 130023, China. hyjin13@mails.jlu.edu.cn.
authors.forename: Hanyong
authors.initials: H
authors.lastname: Jin
authors.affiliation: State Key Laboratory of Theoretical and Computational Chemistry, Institute of Theoretical Chemistry, Jilin University, Changchun 130023, China. guanss12@mails.jlu.edu.cn.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Guan
authors.affiliation: Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, Jilin University, Changchun 130023, China. weiweihan@jlu.edu.cn.
authors.forename: Weiwei
authors.initials: W
authors.lastname: Han
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D048013:Acetivibrio thermocellus / Q000201:enzymology; D016208:Databases, Factual; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000378:metabolism*; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011789:Quantum Theory; D045857:Ruminococcus / Q000201:enzymology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013816:Thermodynamics; D014990:Xylans / Q000378:metabolism
chemical_list: D014990:Xylans; C085118:arabinoxylan; D043364:Endo-1,4-beta Xylanases