title: Lysozyme requires fluctuation of the active site for the manifestation of activity.
abstract: Mutations around His15 which lie far away from the active site, stimulated glycol chitin activity of lysozyme at physiological temperature. Del-Arg14His15 lysozyme, a mutant lysozyme whose Arg14 and His15 were deleted together, and has the highest activity among these mutant lysozymes, had a similar binding ability to a trimer of N-acetyl-glucosamine, a substrate analogue, relative to native lysozyme. This suggests that the increased activity was due to an increased kcat in the catalysis reaction. The H-D exchange rate of the N-1 proton in the Trp63 which is located in the active site cleft, was enhanced in the Del-Arg14His15 lysozyme, while 2-D proton NMR analysis revealed no conformational change around Trp63. We conclude that some sort of fluctuation at the active site might be required for the manifestation of activity. This theory is supported by the finding that the Del-Arg14His15 lysozyme showed a shift in temperature dependency of activity to lower temperatures compared with that of native lysozyme.
authors.affiliation: Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Imoto
authors.forename: T
authors.initials: T
authors.lastname: Ueda
authors.forename: T
authors.initials: T
authors.lastname: Tamura
authors.forename: Y
authors.initials: Y
authors.lastname: Isakari
authors.forename: Y
authors.initials: Y
authors.lastname: Abe
authors.forename: M
authors.initials: M
authors.lastname: Inoue
authors.forename: T
authors.initials: T
authors.lastname: Miki
authors.forename: K
authors.initials: K
authors.lastname: Kawano
authors.forename: H
authors.initials: H
authors.lastname: Yamada
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism; D001120:Arginine; D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D003198:Computer Simulation; D017353:Gene Deletion; D006639:Histidine; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009113:Muramidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013329:Structure-Activity Relationship; D013696:Temperature
chemical_list: D006639:Histidine; D001120:Arginine; D009113:Muramidase; D000117:Acetylglucosamine