Light activates reduction of methotrexate by NADPH in the ternary complex with Escherichia coli dihydrofolate reductase

Yong Qing Chen, Miriam Gulotta, H. T Andrew Cheung, Robert Callender

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Methotrexate (MTX), a strong inhibitor of dihydrofolate reductase (DHFR), has been widely used for chemotherapy for many types of cancer as well as for juvenile rheumatoid arthritis. It mimics folate substrates and binds tightly to the active site of DHFR, perhaps in a conformation close to the transition state of the folate catalyzed reaction. Absorption, fluorescence and ultrasensitive Raman difference spectroscopies show that light-activated MTX reacts with NADPH in the enzyme active site, producing 5,8-dihydromethotrexate (5,8-dihydro-MTX) and NADP+. The reaction, which proceeds with a hydride transfer between C4 (pro-R side) of the nicotinamide ring and N5 of the pteridine ring, is similar to that between folate and NADPH except that the hydride is transferred to C6 in this case. Hence, MTX is catalytically competent in its excited state. Most experiments were performed on the Escherichia coli enzyme, but preliminary studies show that the reaction also occurs with human DHFR.

Original languageEnglish (US)
Pages (from-to)77-85
Number of pages9
JournalPhotochemistry and Photobiology
Volume69
Issue number1
StatePublished - Jan 1999

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Tetrahydrofolate Dehydrogenase
Escherichia
NADP
Methotrexate
Escherichia coli
Folic Acid
Light
hydrides
enzymes
Hydrides
nicotinamide
arthritis
Catalytic Domain
rings
Pteridines
chemotherapy
Complement C4
Folic Acid Antagonists
inhibitors
Chemotherapy

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biophysics

Cite this

Light activates reduction of methotrexate by NADPH in the ternary complex with Escherichia coli dihydrofolate reductase. / Chen, Yong Qing; Gulotta, Miriam; Cheung, H. T Andrew; Callender, Robert.

In: Photochemistry and Photobiology, Vol. 69, No. 1, 01.1999, p. 77-85.

Research output: Contribution to journalArticle

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AB - Methotrexate (MTX), a strong inhibitor of dihydrofolate reductase (DHFR), has been widely used for chemotherapy for many types of cancer as well as for juvenile rheumatoid arthritis. It mimics folate substrates and binds tightly to the active site of DHFR, perhaps in a conformation close to the transition state of the folate catalyzed reaction. Absorption, fluorescence and ultrasensitive Raman difference spectroscopies show that light-activated MTX reacts with NADPH in the enzyme active site, producing 5,8-dihydromethotrexate (5,8-dihydro-MTX) and NADP+. The reaction, which proceeds with a hydride transfer between C4 (pro-R side) of the nicotinamide ring and N5 of the pteridine ring, is similar to that between folate and NADPH except that the hydride is transferred to C6 in this case. Hence, MTX is catalytically competent in its excited state. Most experiments were performed on the Escherichia coli enzyme, but preliminary studies show that the reaction also occurs with human DHFR.

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