Purification of tetrahydrofolate dehydrogenase by affinity chromatography

  • Gauldie J
  • Hillcoat B
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Abstract

Tetrahydrofolate dehydrogenase (5,6,7,8-tetrahydrofolate:NADP+oxidoreductase, EC 1.5.1.3; formerly known as dihydrofolate reductase) from a subline of the L1210 lymphoma has been purified to apparent homogeneity with high recovery by simple steps using agarose to which the enzyme inhibitor amethopterin was coupled. A crude (NH4)2SO4precipitate was applied to a column of methotrexate-agarose which was washed with buffer containing NADPH to increase the binding of the enzyme to the column, and to elute non-enzyme protein. Buffer at high pH, containing dihydrofolate and high concentration of salt then eluted the enzyme together with some nucleic acid contaminants which were removed on a column of Sephadex G-75. © 1972.

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Authors

  • J. Gauldie

  • B. L. Hillcoat

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