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Title 

Cloning, analysis, and expression of the gene for thermostable polyphosphate kinase of Thermus caldophilus GK24 and properties of the recombinant enzyme

Authors 

H S HoeS K LeeDae Sil LeeS T Kwon

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

2003

Citation 

Journal of Microbiology and Biotechnology, vol. 13, no. 1, pp. 139-145

Keywords 

gene cloninggene expressionpolyphosphate kinaseTca PPKthermus caldophilus GK24recombinant enzymeenzyme activityenzyme purificationenzyme subunit

Abstract 

The gene encoding Thermus caldophilus GK24 polyphosphate kinase (Tca PPK) was cloned and sequenced. The gene contains an open reading frame encoding 608 amino acids with a calculated molecular mass of 69,850 Da. The deduced amino acid sequence of Tca PPK showed a 40% homology to Escherichia coli PPK, and 39% to Klebsiella aerogenes PPK. The Tca ppk gene was expressed under the control of the T7lac promoter on pET-22b(+) in E. coli and its enzyme was purified about 70-fold with 36% yield, following heating and HiTrap chelating HP column chromatography. The native enzyme was found to have an approximate molecular mass of 580,000 Da and consisted of eight subunits. The optimum pH and temperature of the enzyme were 5.5 and 70°C, respectively. A divalent cation was required for the enzyme activity, with Mg2+ being the most effective.

ISSN 

1017-7825

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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