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The Enzyme that Reduces Oxidized Cytoglobin in Bovine Liver: An Exploration

Nabilla Sonia Sahara 1, Mohamad Sadikin 2, and Sri Widia A. Jusman 3
1. Master Program in Biomedical Sciences, Faculty of Medicine, University Indonesia, Jakarta, Indonesia
2. Department of Biochemistry & Molecular Biology, Faculty of Medicine, University Indonesia, Jakarta, Indonesia
3. Center of Hypoxia & Oxidative Stres Studies, Faculty of Medicine University Indonesia, Jakarta, Indonesia
Abstract—Oxidized cytoglobin (Cygb) can be reduced by supernatant of bovine liver cell homogenate as demonstrated by increased production of Cygb-Fe2+ from Cygb-Fe3+. We hypothesized that, in bovine liver tissue, there is a protein that acts as reductase, which is analogous to diaphorase, acting on methemoglobin (metHb), and analogous to cytochrome b5 reductase 3 (CYB5R3), acting on metmyoglobin (metMb). The aim of this study is to explore the enzyme that can reduce oxidized Cygb to reduced Cygb. The putative enzyme in bovine liver homogenate was isolated using RIPA lysis buffer, purified by Cibacron blue chromatography, and confirmed by SDS PAGE and western blot. The activity of the suspect reductase enzyme was determined by the ratio of maximum absorbance between Cygb-Fe3+ (metCygb) and Cygb-Fe2+ (deoxyCygb). We discovered that the reducing capacity of fraction, purified by Cibacron blue chromatography, was weaker than the cell extracted from bovine liver. The gel electrophoresis analysis indicated that the Cibacron blue fraction molecular weight was ~50 and ~60 kDa, whereas CYB5R3 was 34 kDa. Moreover, metcytoglobin could not be reduced by diaphorase. We concluded, there is a reductase enzyme in the cell extract that can reduce Cygb-Fe3+ to Cygb-Fe2+; however, it is neither diaphorase nor CYB5R3.
 
Index Terms—chromatography, cytoglobin, purification, enzyme, reductase

Cite: Nabilla Sonia Sahara, Mohamad Sadikin, and Sri Widia A. Jusman, "The Enzyme that Reduces Oxidized Cytoglobin in Bovine Liver: An Exploration," International Journal of Pharma Medicine and Biological Sciences, Vol. 9, No. 3, pp. 129-133, July 2020. doi: 10.18178/ijpmbs.9.3.129-133

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