Expression, purification, and characterization of the oligomeric states of rubredoxin oxidoreductase from Methanobacterium thermoautotrophicum.
The rubredoxin oxidoreductase from Methanobacterium thermoautotrophicum was successfully overexpressed in Escherichia coli with 6x His-Tag and purified by using Ni-NTA technology. It was characterized by SDS and native polyacrylamide gel electrophoresis (PAGE), as well as size-exclusion chromatography. The protein was pure, judged by SDS-PAGE, but three or more oligomeric species were observed by native PAGE and size-exclusion chromatography. The smallest rubredoxin oxidoreductase species is the dimer. The multiple species are stable and remain in their respective oligomeric states, judged by the chromatographic and electrophoretic results. A model is proposed in order to explain the structural basis for these results.
Chae
Young Kee
YK
Department of Applied Chemistry and Recombinant Protein Expression Center, Sejong University, Seoul, Korea. ykchae@sejong.ac.kr
D000595:Amino Acid Sequence; D019281:Dimerization; D017014:Methanobacterium / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D011108:Polymers / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment
D011108:Polymers; C023091:rubredoxin-NAD+ reductase; D009247:NADH, NADPH Oxidoreductases