Crystal structure of Tapes japonica lysozyme with substrate analogue: Structural basis of the catalytic mechanism and manifestation of its chitinase activity accompanied by quaternary structural change

Takashi Goto, Yoshito Abe, Yoshimitsu Kakuta, Kohei Takeshita, Taiji Imoto, Tadashi Ueda

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Abstract

Tapes japonica lysozyme (TJL) is classified as a member of the recently established i-type lysozyme family. In this study, we solved the crystal structure of TJL complexed with a trimer of N-acetylglucosamine to 1.6Å resolution. Based on structure and mutation analyses, we demonstrated that Glu-18 and Asp-30 are the catalytic residues of TJL. Furthermore, the present findings suggest that the catalytic mechanism of TJL is a retaining mechanism that proceeds through a covalent sugar-enzyme intermediate. On the other hand, the quaternary structure in the crystal revealed a dimer formed by the electrostatic interactions of catalytic residues (Glu-18 and Asp-30) in one molecule with the positive residues at the C terminus in helix 6 of the other molecule. Gel chromatography analysis revealed that the TJL dimer remained intact under low salt conditions but that it dissociated to TJL monomers under high salt conditions. With increasing salt concentrations, the chitinase activity of TJL dramatically increased. Therefore, this study provides novel evidence that the lysozyme activity of TJL is modulated by its quaternary structure.

Original languageEnglish
Pages (from-to)27459-27467
Number of pages9
JournalJournal of Biological Chemistry
Volume282
Issue number37
DOIs
Publication statusPublished - Sep 14 2007

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Chitinases
Muramidase
Tapes
Crystal structure
Substrates
Salts
Dimers
Molecules
Chromatography
Coulomb interactions
Static Electricity
Sugars
Gel Chromatography
Monomers
Gels

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

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title = "Crystal structure of Tapes japonica lysozyme with substrate analogue: Structural basis of the catalytic mechanism and manifestation of its chitinase activity accompanied by quaternary structural change",
abstract = "Tapes japonica lysozyme (TJL) is classified as a member of the recently established i-type lysozyme family. In this study, we solved the crystal structure of TJL complexed with a trimer of N-acetylglucosamine to 1.6{\AA} resolution. Based on structure and mutation analyses, we demonstrated that Glu-18 and Asp-30 are the catalytic residues of TJL. Furthermore, the present findings suggest that the catalytic mechanism of TJL is a retaining mechanism that proceeds through a covalent sugar-enzyme intermediate. On the other hand, the quaternary structure in the crystal revealed a dimer formed by the electrostatic interactions of catalytic residues (Glu-18 and Asp-30) in one molecule with the positive residues at the C terminus in helix 6 of the other molecule. Gel chromatography analysis revealed that the TJL dimer remained intact under low salt conditions but that it dissociated to TJL monomers under high salt conditions. With increasing salt concentrations, the chitinase activity of TJL dramatically increased. Therefore, this study provides novel evidence that the lysozyme activity of TJL is modulated by its quaternary structure.",
author = "Takashi Goto and Yoshito Abe and Yoshimitsu Kakuta and Kohei Takeshita and Taiji Imoto and Tadashi Ueda",
year = "2007",
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T1 - Crystal structure of Tapes japonica lysozyme with substrate analogue

T2 - Structural basis of the catalytic mechanism and manifestation of its chitinase activity accompanied by quaternary structural change

AU - Goto, Takashi

AU - Abe, Yoshito

AU - Kakuta, Yoshimitsu

AU - Takeshita, Kohei

AU - Imoto, Taiji

AU - Ueda, Tadashi

PY - 2007/9/14

Y1 - 2007/9/14

N2 - Tapes japonica lysozyme (TJL) is classified as a member of the recently established i-type lysozyme family. In this study, we solved the crystal structure of TJL complexed with a trimer of N-acetylglucosamine to 1.6Å resolution. Based on structure and mutation analyses, we demonstrated that Glu-18 and Asp-30 are the catalytic residues of TJL. Furthermore, the present findings suggest that the catalytic mechanism of TJL is a retaining mechanism that proceeds through a covalent sugar-enzyme intermediate. On the other hand, the quaternary structure in the crystal revealed a dimer formed by the electrostatic interactions of catalytic residues (Glu-18 and Asp-30) in one molecule with the positive residues at the C terminus in helix 6 of the other molecule. Gel chromatography analysis revealed that the TJL dimer remained intact under low salt conditions but that it dissociated to TJL monomers under high salt conditions. With increasing salt concentrations, the chitinase activity of TJL dramatically increased. Therefore, this study provides novel evidence that the lysozyme activity of TJL is modulated by its quaternary structure.

AB - Tapes japonica lysozyme (TJL) is classified as a member of the recently established i-type lysozyme family. In this study, we solved the crystal structure of TJL complexed with a trimer of N-acetylglucosamine to 1.6Å resolution. Based on structure and mutation analyses, we demonstrated that Glu-18 and Asp-30 are the catalytic residues of TJL. Furthermore, the present findings suggest that the catalytic mechanism of TJL is a retaining mechanism that proceeds through a covalent sugar-enzyme intermediate. On the other hand, the quaternary structure in the crystal revealed a dimer formed by the electrostatic interactions of catalytic residues (Glu-18 and Asp-30) in one molecule with the positive residues at the C terminus in helix 6 of the other molecule. Gel chromatography analysis revealed that the TJL dimer remained intact under low salt conditions but that it dissociated to TJL monomers under high salt conditions. With increasing salt concentrations, the chitinase activity of TJL dramatically increased. Therefore, this study provides novel evidence that the lysozyme activity of TJL is modulated by its quaternary structure.

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