title: Zinc in lipase L1 from Geobacillus stearothermophilus L1 and structural implications on thermal stability.
abstract: Lipase L1 from Geobacillus stearothermophilus L1 contains an unusual extra domain, making a tight intramolecular interaction with the main catalytic domain through a Zn2+-binding coordination. To elucidate the role of the Zn2+, we disrupted the Zn2+-binding site by mutating the zinc-ligand residues (H87A, D61A/H87A, and D61A/H81A/H87A/D238A). The activity vs. temperature profiles of the mutant enzymes showed that the disruption of the Zn2+-binding site resulted in a notable decrease in the optimal temperature for maximal activity from 60 to 45-50 degrees C. The mutations also abolished the Zn2+-induced thermal stabilization. The wild-type enzyme revealed a 34.6-fold increase in stabilization with the addition of Zn2+ at 60 degrees C, whereas the mutant enzymes exhibited no response to Zn2+. Additional circular dichroism spectroscopy studies also confirmed the structural stabilizing role of Zn2+ on lipase L1 at elevated temperatures.
authors.affiliation: Laboratory of Microbial Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB),Yusong, Daejon 305-600, Korea.
authors.forename: Won-Chan
authors.initials: WC
authors.lastname: Choi
authors.forename: Myung Hee
authors.initials: MH
authors.lastname: Kim
authors.forename: Hyeon-Su
authors.initials: HS
authors.lastname: Ro
authors.forename: Sang Ryeol
authors.initials: SR
authors.lastname: Ryu
authors.forename: Tae-Kwang
authors.initials: TK
authors.lastname: Oh
authors.forename: Jung-Kee
authors.initials: JK
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D001406:Bacillaceae / Q000201:enzymology*; D001665:Binding Sites / Q000235:genetics; D020134:Catalytic Domain; D004795:Enzyme Stability; D008049:Lipase / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary; D013696:Temperature; D015032:Zinc / Q000737:chemistry*
chemical_list: D008049:Lipase; D015032:Zinc