title: Residue Tyr224 is critical for the thermostability of Geobacillus sp. RD-2 lipase.
abstract: A thermophilic lipase (lipGRD) from Geobacillus sp. RD-2, isolated from a hot spring in Yunnan, China, was cloned and over-expressed in Escherichia coli. The function of the conserved residue, Tyr224, near the presumed temperature switch site was analyzed by site-directed saturation mutagenesis. The activity of the wild type lipGRD was optimal at 55 degrees C and pH 7.5, but that from mutant Y224C was optimally active at 35 degrees C, whereas Y224P lipase was optimally active at 65 degrees C. Furthermore, the latter lipase retained 60% of its activity after incubation at 65 degrees C for 5 h. The conserved residue Tyr224, which is close to the lid helix, is the key amino acid residue determining the thermostability of the thermostable lipase.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, 27 Shanda Nan Rd., 250100, Jinan, People's Republic of China.
authors.forename: Lei
authors.initials: L
authors.lastname: Wu
authors.forename: Bo
authors.initials: B
authors.lastname: Liu
authors.forename: Ye
authors.initials: Y
authors.lastname: Hong
authors.forename: Duohong
authors.initials: D
authors.lastname: Sheng
authors.forename: Yulong
authors.initials: Y
authors.lastname: Shen
authors.forename: Jinfeng
authors.initials: J
authors.lastname: Ni
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D003001:Cloning, Molecular; D004795:Enzyme Stability / Q000235:genetics / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000235:genetics; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000235:genetics / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D014443:Tyrosine; D008049:Lipase; C428386:thermostable lipase