Identification of a novel gentisate 1,2-dioxygenase from <Emphasis Type="Italic">Silicibacter pomeroyi</Emphasis> |
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Authors: | Dongqi Liu Tingting Zhu Li Fan Junming Quan Hongchun Guo Jinren Ni |
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Institution: | College of Environmental Sciences, Peking University, Beijing, 100871, P.R. China. liudongqi@iee.pku.edu.cn |
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Abstract: | A 1,125-bp long ORF encoding a novel gentisate 1,2-dioxygenase with two-domain bicupins was cloned from Silicibacter pomeroyi DSS-3 and expressed in Escherichia coli. The resulting product was purified to homogeneity and partially characterized. Non-reductive SDS-PAGE and gel filtration showed that the active recombinant gentisate 1,2-dioxygenase had an estimated molecular mass of 132 kDa, and reductive SDS-PAGE indicated an approximate size of 45 kDa. The enzyme thus appears to be a homotrimeric protein. This is in contrast to the homotetrameric or dimeric protein of the gentisate 1,2-dioxygenases that have been characterized thus far. The K (m) and K (cat)/K (m) for gentisate were 12 muM and 653 x 10(4) M(-1 )s(-1); the pI was 4.6-4.8. It was optimally active at 40 degrees C and pH 8.0. |
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Keywords: | Gentisate 1 2-dioxygenase Purification Recombinant Silicibacter pomeroyi |
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