Large scale production and characterisations of dihydrofolate reductase from a methotrexate-resistant human lymphoid cell line |
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| Authors: | Aftab Alam |
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| Institution: | (1) Astbury Department of Biophysics, University of Leeds, LS2 9JT Leeds, England;(2) BioKalfs-research and development, Suite 8, 2nd Floor, Royal House, 28 Sovereign Street, LS1 4BJ Leeds, England |
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| Abstract: | Dihydrofolate reductase has been purified from a methotrexate-resistant human lymphoid cell line (CCRF/CEM-R3) and up to 1
mg of enzyme has been obtained from 5 litres of culture. The enzyme has a molecular weight of 22000 ±500 as determined by
gel filtration. The pH activity profile shows a single optimum at pH 7.7, where marked activation is observed by addition
of 0.2 M NaCl. TheK
m for NADPH is 3μM and dihydrofolate 0.7μM. The binding constant for the inhibitor, methotrexate, is 29 pM |
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| Keywords: | Human dihydrofolate reductase purification characterization |
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