title: Ancestral amino acid substitution improves the thermal stability of recombinant lignin-peroxidase from white-rot fungi, Phanerochaete chrysosporium strain UAMH 3641.
abstract: Stabilizing enzymes from mesophiles of industrial interest is one of the greatest challenges of protein engineering. The ancestral mutation method, which introduces inferred ancestral residues into a target enzyme, has previously been developed and used to improve the thermostability of thermophilic enzymes. In this report, we studied the ancestral mutation method to improve the chemical and thermal stabilities of Phanerochaete chrysosporium lignin peroxidase (LiP), a mesophilic fungal enzyme. A fungal ancestral LiP sequence was inferred using a phylogenetic tree comprising Basidiomycota and Ascomycota fungal peroxidase sequences. Eleven mutant enzymes containing ancestral residues were designed, heterologously expressed in Escherichia coli and purified. Several of these ancestral mutants showed higher thermal stabilities and increased specific activities and/or kcat/KM than those of wild-type LiP.
authors: Department of Applied Biology, Faculty of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1, Horinouchi, Hachioji, Tokyo 192-0392, Japan.
authors: Yasuyuki
authors: Y
authors: Semba
authors: Department of Applied Biology, Faculty of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1, Horinouchi, Hachioji, Tokyo 192-0392, Japan Top Runner Incubation Center for Academia-Industry Fusion, Department of Bioengineering, Faculty of Engineering, Nagaoka University of Technology, 1603-1, Kamitomiokamachi, Nagaoka, Niigata 940-2188, Japan.
authors: Manabu
authors: M
authors: Ishida
authors: Department of Applied Biology, Faculty of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1, Horinouchi, Hachioji, Tokyo 192-0392, Japan.
authors: Shin-ichi
authors: S
authors: Yokobori
authors: Department of Applied Biology, Faculty of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1, Horinouchi, Hachioji, Tokyo 192-0392, Japan yamagish@toyaku.ac.jp yamagish@ls.toyaku.ac.jp.
authors: Akihiko
authors: A
authors: Yamagishi
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D003001:Cloning, Molecular; D004795:Enzyme Stability / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D020075:Phanerochaete / Q000201:enzymology*; D010802:Phylogeny; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D013696:Temperature; D044366:Transition Temperature
chemical_list: D011994:Recombinant Proteins; D010544:Peroxidases; C042858:lignin peroxidase
keywords: Phanerochaete chrysosporium; ancestral mutation method; lignin peroxidase; thermostabilization