title: Engineering surface hydrophobicity improves activity of Bacillus thermocatenulatus lipase 2 enzyme.
abstract: Bacillus thermocatenulatus lipase 2 (BTL2) is a promising industrial enzyme used in biodiesel production. Although BTL2 has high thermostability and good resistance to organic solvents, the activity of BTL2 is suboptimal for industrial processes. To improve BTL2 activity, we engineered BTL2 lipase by modulating hydrophobicity of its lid domain. Through site-directed mutagenesis, we constructed three mutants, namely Y225F+S232A, S232A+T236V and Q185L, to cover all uncharged hydrophilic amino acids within the lid domain. Activities of these mutants were characterized. Our findings suggest that one mutant (Y225F+S232A) showed ∼35% activity increase in catalyzing heterogeneous hydrolytic reactions relevant for industrial applications. A mathematical framework was established to account for different molecular events that contribute to the observed apparent catalytic activities. Increases in hydrophobicity of lid domains were associated with increased interfacial adsorption of lipases and lower molecular enzymatic activities. The measured apparent activities of lipases include contributions from both events. Lid hydrophobicity can thus result in different changes in lipase activities depending on the mutation site. Our work demonstrates the feasibility of increasing BTL2 activity by modulating the hydrophobicity of lid domains and provides some guidelines for further improving BTL2 activity.
authors.affiliation: Department of Chemical Engineering, Institute of Applied Chemistry, Tsinghua University, Beijing, China.
authors.forename: Ting
authors.initials: T
authors.lastname: Tang
authors.affiliation: School of Chemical Engineering, Purdue University, West Lafayette, IN, USA.
authors.forename: Chongli
authors.initials: C
authors.lastname: Yuan
authors.affiliation: School of Chemical Engineering, Purdue University, West Lafayette, IN, USA.
authors.forename: Hyun-Tae
authors.initials: HT
authors.lastname: Hwang
authors.affiliation: Department of Chemical Engineering, Institute of Applied Chemistry, Tsinghua University, Beijing, China.
authors.forename: Xuebing
authors.initials: X
authors.lastname: Zhao
authors.affiliation: School of Chemical Engineering, Purdue University, West Lafayette, IN, USA.
authors.forename: Doraiswami
authors.initials: D
authors.lastname: Ramkrishna
authors.affiliation: Department of Chemical Engineering, Institute of Applied Chemistry, Tsinghua University, Beijing, China.
authors.forename: Dehua
authors.initials: D
authors.lastname: Liu
authors.affiliation: School of Chemical Engineering, Purdue University, West Lafayette, IN, USA. avarma@purdue.edu.
authors.forename: Arvind
authors.initials: A
authors.lastname: Varma
mesh_terms: D001407:Bacillus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D013499:Surface Properties
chemical_list: D001426:Bacterial Proteins; D008049:Lipase
keywords: Bacillus thermocatenulatus lipase 2 (BTL2); Heterogeneous reaction; Hydrophobicity; Interfacial adsorption; Lid