Biochemical characterization of glyceraldehyde-3-phosphate dehydrogenase from <Emphasis Type="Italic">Thermococcus kodakarensis</Emphasis> KOD1 |
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Authors: | Baolei Jia Le Thuy Linh Sangmin Lee Bang Phuong Pham Jinliang Liu Hongyu Pan Shihong Zhang Gang-Won Cheong |
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Institution: | (1) College of Plant Sciences, Jilin University, Changchun, 130-062, China;(2) Division of Applied Life Sciences (BK21 Program), Gyeongsang National University, Jinju, 660-701, Korea;(3) Environmental Biotechnology National Core Research Center, Gyeongsang National University, Jinju, 660-701, Korea; |
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Abstract: | Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) plays an essential role in glycolysis by catalyzing the conversion of d-glyceraldehyde 3-phosphate (d-G3P) to 1,3-diphosphoglycerate using NAD+ as a cofactor. In this report, the GAPDH gene from the hyperthermophilic archaeon Thermococcus kodakarensis KOD1 (GAPDH-tk) was cloned and the protein was purified to homogeneity. GAPDH-tk exists as a homotetramer with a native molecular
mass of 145 kDa; the subunit molecular mass was 37 kDa. GAPDH-tk is a thermostable protein with a half-life of 5 h at 80–90°C.
The apparent K
m values for NAD+ and d-G3P were 77.8 ± 7.5 μM and 49.3 ± 3.0 μM, respectively, with V
max values of 45.1 ± 0.8 U/mg and 59.6 ± 1.3 U/mg, respectively. Transmission electron microscopy (TEM) and image processing
confirmed that GAPDH-tk has a tetrameric structure. Interestingly, GAPDH-tk migrates as high molecular mass forms (~232 kDa
and ~669 kDa) in response to oxidative stress. |
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Keywords: | |
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