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- Purification and Characterization of a Catalase from Photosynthetic Bacterium Rhodospirillum rubrum S1 Grown under Anaerobic Conditions
- Yoon-Suk Kang , Dong-Heon Lee , Byoung-Jun Yoon , Duck-Chul Oh
- J. Microbiol. 2006;44(2):185-191.
- DOI: https://doi.org/2366 [pii]
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Abstract
- The photosynthetic bacterium, Rhodospirillum rubrum S1, when grown under anaerobic conditions, generated three different types of catalases. In this study, we purified and characterized the highest molecular weight catalase from the three catalases. The total specific catalase activity of the crude cell extracts was 88 U/mg. After the completion of the final purification step, the specific activity of the purified catalase was 1,256 U/mg. The purified catalase evidenced an estimated molecular mass of 318 kDa, consisting of four identical subunits, each of 79 kDa. The purified enzyme exhibited an apparent Km value of 30.4 mM and a Vmax of 2,564 U against hydrogen peroxide. The enzyme also exhibited a broad optimal pH (5.0~9.0), and remained stable over a broad temperature range (20°C~60°C). It maintained 90% activity against organic solvents (ethanol/chloroform) known hydroperoxidase inhibitors, and exhibited no detectable peroxidase activity. The catalase activity of the purified enzyme was reduced to 19% of full activity as the result of the administration of 10 mM 3-amino-1,2,4-triazole, a heme-containing catalase inhibitor. Sodium cyanide, sodium azide, and hydroxylamine, all of which are known heme protein inhibitors, inhibited catalase activity by 50% at concentrations of 11.5 μM, 0.52 μM, and 0.11 μM, respectively. In accordance with these findings, the enzyme was identified as a type of monofunctional catalase.
- Analysis of Catalases from Photosynthetic Bacterium Rhodospirillum rubrum S1
- Hee-Kyoung Lim , Young-Mi Kim , Dong-Heon Lee , Hyung-Yeel Kahng , Duck-Chul Oh
- J. Microbiol. 2001;39(3):168-176.
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Abstract
- Five different types of catalases from photosynthetic bacterium Rhodospirillum rubrum S1 grown aerobically in the dark were found in this study, and designated Cat1 (350 kDa), Cat2 (323 kDa), Cat3 (266 kDa), Cat4 (246 kDa), and Cat5 (238 kDa). Analysis of native PAGE revealed that Cat2, Cat3, and Cat4 were also produced in the cells anaerobically grown in the light. It is notable that only Cat2 was expressed much more strongly in response to the anaerobic condition. Enzyme activity staining demonstrated that Cat3 and Cat4 had bifunctional catalase-peroxidase activities, while Cat1, Cat2, and Cat5 were typical monofunctional catalases. S1 cells grown aerobically in the presence of malate as the sole source of carbon exhibited an apparent catalase Km value of 10 mM and a Vmax of about 705 U/mg protein at late stationary growth phase. The catalase activity of S1 cells grown in the anaerobic environment exhibited a much lower Vmax of about 109 U/mg protein at late logarithmic growth phase. The catalytic activity was stable in the broad range of temperatures (30 C-60 C), and pH (6.0-10.0). R. rubrum S1 was much more resistant to H_2 O_2 in the stationary growth phase than in the exponential growth phase regardless of growth conditions. Cells of stationary growth phase treated with 15 mM H 2 O 2 for 1 h showed 3-fold higher catalase activities than the untreated cells. In addition, L-glutamate induced an 80-fold increase in total catalase activity of R. rubrum S1 compared with malic acid. Through fraction analyses of S1 cells, Cat2, Cat3, Cat4 and Cat5 were found in both cytoplasm and periplasm, while Cat1 was localized only in the cytoplasm.
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