Reconstitution and function of Tetragenococcus halophila chaperonin 60 tetradecamer.
Tetragenococcus halophila originally isolated from soy sauce is a halophilic lactic acid bacterium which can grow under 4 M sodium chloride. T. halophila chaperonin composed of a core moiety of chaperonin 60 (cpn60) and a lid moiety of chaperonin 10 (cpn10), is thought to contribute to host halotolerant capability. In this report, we reconstituted and characterized the core complex of T. halophila chaperonin by using a recombinant T. halophila cpn60 (Tcpn60) overexpressed in Escherichia coli. The reconstitution of Tcpn60 was performed in the presence of 10 mM MgCl2, 2 mM ATP and 0.8 M (NH4)2SO4 and the resultant oligomer was purified by gel filtration chromatography. Electron microscopy of the reconstituted Tcpn60 revealed a double toroidal tetradecameric structure that is characteristic of bacterial cpn60. The T. halophila tetradecamer cpn60 exhibited an ATPase activity and a refolding activity of both chemically and thermally denatured enolases under wide range of salt concentrations. Furthermore, we demonstrated that heterologous expression of Tcpn60 allowed the normal growth of host Escherichia coli cells under salt stress conditions and this effect was further enhanced by co-expression with Tcpn10. These results suggested that Tcpn60 contributes to the halotolerance property of T. halophila cell as a tetradecamer complex, probably associated with the Tcpn10 complex.
Tosukhowong
Amonlaya
A
Laboratory of Microbial Technology, Division of Microbial Science and Technology, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
Nakayama
Jiro
J
Mizunoe
Yoshimitsu
Y
Sugimoto
Shinya
S
Fukuda
Daisuke
D
Sonomoto
Kenji
K
D049109:Cell Proliferation; D018834:Chaperonin 60 / Q000032:analysis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007778:Lactobacillus / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000032:analysis / Q000737:chemistry / Q000648:ultrastructure; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000648:ultrastructure; D012965:Sodium Chloride / Q000737:chemistry*
D018834:Chaperonin 60; D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D012965:Sodium Chloride