title: Enhancement of methanol resistance of Yarrowia lipolytica lipase 2 using β-cyclodextrin as an additive: Insights from experiments and molecular dynamics simulation.
abstract: The methanol resistance of lipase is a critical parameter in enzymatic biodiesel production. In the present work, the methanol resistance of Yarrowia lipolytica Lipase 2 (YLLIP2) was significantly improved using β-cyclodextrin (β-CD) as an additive. According to the results, YLLIP2 with β-CD exhibited approximately 7000U/mg specific activity in 30wt% methanol for 60min compared with no activity without β-CD under the same conditions. Molecular dynamics (MD) simulation results indicated that the β-CD molecules weakened the conformational change of YLLIP2 and maintained a semi-open state of the lid by overcoming the interference caused by methanol molecules. Furthermore, the β-CD molecule could directly stabilize "pathway" regions (e.g., Asp61-Asp67) and indirectly stabilize "pathway" regions (e.g., Gly44-Phe50) by forming hydrogen bonds with "pathway" regions and nearby "pathway" regions, respectively. The regions stabilized by the β-CD molecule then prevented the closure of active pockets, thus retaining the enzymatic activity of YLLIP2 with β-CD in methanol solvent.
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China.
authors.forename: Hao
authors.initials: H
authors.lastname: Cao
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China.
authors.forename: Haiyang
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China; Amoy-BUCT Industrial of Bio-tech novation Institute, Amoy, 361026, PR China.
authors.forename: Kaili
authors.initials: K
authors.lastname: Nie
authors.affiliation: State Key Laboratory of Chemical Resource Engineering, Institute of Materia Medica, College of Science, Beijing University of Chemical Technology, Beijing, 100029, PR China.
authors.forename: Ming
authors.initials: M
authors.lastname: Lei
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China; Amoy-BUCT Industrial of Bio-tech novation Institute, Amoy, 361026, PR China. Electronic address: dengli@mail.buct.edu.cn.
authors.forename: Li
authors.initials: L
authors.lastname: Deng
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China.
authors.forename: Fang
authors.initials: F
authors.lastname: Wang
authors.affiliation: Beijing Bioprocess Key Laboratory, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, PR China. Electronic address: twtan@mail.buct.edu.cn.
authors.forename: Tianwei
authors.initials: T
authors.lastname: Tan
mesh_terms: D056804:Biofuels; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D004795:Enzyme Stability / Q000187:drug effects; D005656:Fungal Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D008049:Lipase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D000432:Methanol / Q000494:pharmacology*; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation / Q000187:drug effects; D025062:Yarrowia / Q000201:enzymology*; D047392:beta-Cyclodextrins / Q000737:chemistry / Q000494:pharmacology
chemical_list: D056804:Biofuels; D005656:Fungal Proteins; D047392:beta-Cyclodextrins; D002265:Carboxylic Ester Hydrolases; D008049:Lipase; C000606101:YLLIP2 protein, Yarrowia lipolytica; D000432:Methanol
keywords: Lipase; Methanol resistance; Molecular dynamics; Surface modification; β-cyclodextrin