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KMID : 0380020120270060341
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Korean Journal of Biotechnology and Bioengineering
2012 Volume.27 No. 6 p.341 ~ p.346
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Purification and Characterization of an Indican-hydrolyzing β-glucosidase from Agrobacterium tumefaciens
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Hwang Chang-Sun
Lee Jin-Young
Kim Geun-Joong
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Abstract
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Indican (indoxyl-β-D-glucoside) is a colorless natural compound and can be used as a precursor for the production of indigo. This production step only require an enzyme, β-glucosidase, that readily screened from microbial resource by using selective media supplemented with indican as a sole carbon source. Agrobacterium tumefaciens was well grown in this media and thus presumed to produce a related enzyme. The corresponding gene, encoding a protein with a calculated molecular mass of 51 kDa, was cloned and overexpressed as MBP fusion proteins. The purified enzyme was determined to be a dimer and showed the maximum activity for indican at pH 7.0 and 40℃. The kinetic parameters for indican, Km and Vmax, were determined to be 1.4 mM and 373.8 μM/min/mg, respectively. The conversion yield of indican into indigo using this enzyme was about 1.7-1.8 folds higher than that of previously isolated enzyme from Sinorhizobium meliloti. Additionally, this enzyme was able to hydrolyze various β-1,4 glycoside substrates.
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KEYWORD
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Indican, Glucosidase, Purification, Agrobacterium, Indigo
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