• Journal of Plant Biotechnology
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• 제3권1호
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• pp.13-17
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• 2001

A Chromium (VI)[Cr(VI)] reductase gene from heavy metal reducing bacteria Pseudomonas aeruginosa HP014 was used to transform tobacco plant cells. A chimeric construct containing the Cr(VI) reductase gene was transfered to tobacco leaf disks using an Agrobacteriun tumefaciens binary vector system. From the leaf disks, transformed plantlets were regenerated. Hybridization experiments demonstrated that the Cr(VI) reductase gene was inserted into and expressed in the regenerated plants. The Cr(VI) reduction activity showed that the transgenic plants may be a another possible tool to reduce the pollution of the toxic Cr(VI) in soil.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.672-675
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• 2000

E.coli ATCC 33456은 6가 크롬 환원능이 있다고 보고되어진 미생물보다 비교적 높은 활성을 지닌다. E.coli ATCC 33456에서 6가 크롬 환원효소를 클로닝하기 위해서 colony hybridization과 southern hybridization을 통하여 2개의 형질 전환 균주를 획득할 수 있었다. 두 균주 모두 E.coli ATCC 33456이 가진 6가 크롬 환원능보다 약 2.5-4배정도 높은 활성을 보였으며, 두 균주의 조효소액을 가지고 immuno-blotting을 실시한 결과, E.coli ATCC 33456에서 분리 정제된 6가 크롬 환원효소의 항체가 약 42Kda에서 반응하는 것을 확인할 수 있었으며, 이 결과는 분리 정제된 6가 크롬 환원효소와 매우 유사한 유전자 산물이 두 균주에서 나온다는 것을 보여주었다.

• Journal of Microbiology and Biotechnology
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• 제10권5호
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• pp.580-586
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• 2000

A soluble Cr(VI) reductase was purified from the Cr(VI) reducing strain Escherichia coli ATCC33456 by ammonium sulfate fractionation, and chromatographies on Q-Sepharose FF, Cibacron blue 3GA dye affinity, Mono-Q 5/5, and Superdex 200 HR 10/30 columns. The estimated molecular mass of the purified enzyme was 27 kDa on SDS-polyacrylamide gel electrophoresis and 54 kDa on gel filtration, thus indicating a dimeric structure. The isoelectric point of the enzyme was pH 4.85. The optimum reaction pH and storage pH were both 7.0, the optimum reaction temperature was $37^{\circ}C$, and the storage temperature was $4^{\circ}C$. NADH and NADPH both served as electron donors for the reductase, with $V_{max}$ of 68.3 ${\mu}M$ Cr(VI)/min/mg protein and Km of 7.6 $\mu$M using HADH, and Vmax of 42.3 ${\mu}M$ Cr(VI)/min/mg protein and Km of 14.6 $\muM$ using NADPH. When 1 mM EDTA was added, the Cr(VI) reducing activity increased 4-fold.

• Journal of Microbiology
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• 제43권1호
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• pp.21-27
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• 2005

A soluble Cr(VI) reductase was purified from the cytoplasm of Escherichia coli ATCC 33456. The molecular mass was estimated to be 84 and 42 kDa by gel filtration and SDS-polyacrylamide gel electrophoresis, respectively, indicating a dimeric structure. The pI was 4.66, and optimal enzyme activity was obtained at pH 6.5 and $37^{\circ}C$. The most stable condition existed at pH 7.0. The purified enzyme used both NADPH and NADH as electron donors for Cr(VI) reduction, while NADPH was the better, conferring 61% higher activity than NADH. The $K_m$ values for NADPH and NADH were determined to be 47.5 and 17.2 umol, and the $V_max$ values 322.2 and 130.7 umol Cr(VI) $min^{-1}mg^{-1}$ protein, respectively. The activity was strongly inhibited by N-ethylmalemide, $Ag^{2+},\;Cd^{2+},\;Hg^{2+}$, and $Zn^{2+}$. The antibody against the enzyme showed no immunological cross reaction with those of other Cr(VI) reducing strains.

• Journal of Microbiology and Biotechnology
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• 제22권4호
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• pp.547-554
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• 2012

Chromium is generated from several industrial processes. It occurs in different oxidation states, but Cr(III) and Cr(VI) are the most common ones. Cr(VI) is a toxic, soluble environmental contaminant. Some bacteria are able to reduce hexavalent chromium to the insoluble and less toxic Cr(III), and thus chromate bioremediation is of considerable interest. An indigenous chromium-reducing bacterial strain, Rb-2, isolated from a tannery water sample, was identified as Ochrobactrum intermedium, on the basis of 16S rRNA gene sequencing. The influence of factors like temperature of incubation, initial concentration of Cr, mobility of bacteria, and different carbon sources were studied to test the ability of the bacterium to reduce Cr(VI) under variable environmental conditions. The ability of the bacterial strain to reduce hexavalent chromium in artificial and industrial sewage water was evaluated. It was observed that the mechanism of resistance to metal was not due to the change in the permeability barrier of the cell membrane, and the enzyme activity was found to be inductive. Intracellular reduction of Cr(VI) was proven by reductase assay using cell-free extract. Scanning electron microscopy revealed chromium precipitates on bacterial cell surfaces, and transmission electron microscopy showed the outer as well as inner distribution of Cr(VI). This bacterial strain can be useful for Cr(VI) detoxification under a wide range of environmental conditions.