Purification and characterization of a mitochondrial DNA polymerase from cultured tobacco cells

Sato, K.; Fukuda, H.

Purification and characterization of a mitochondrial DNA polymerase from cultured tobacco cells

1996

Sato, K. | Fukuda, H.

The mitochondrial DNA polymerase of tobacco BY-2 cells was purified more than 1,200-fold by column chromatography on DEAE cellulose, phosphocellulose, and AffiPrep heparin. The enzyme was classified as a ~type DNA polymerase, in view of the inhibition of its activity by N-ethylmaleimide and dideoxy TTP, the absence of inhibition by aphidicolin and arabinosyl CTP, the stimulation by KCl, and the ability of the enzyme to utilize poly(A)(dT),2 ,' in the presence of Mn2+ ions. The molecular mass of the native mitochondrial DNA polymerase was estimated to be 70-110 kDa by column chromatography on Superose 12. When DNA polymerase activity was analyzed after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the activity that polymerized DNA was observed as a single band of a protein with a molecular mass of approximately 110 kDa. Therefore, the tobacco mitochondrial DNA polymerase appears to consist of a single subunit of 110 kDa.

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