PTR1: A Reductase Mediating Salvage of Oxidized Pteridines and Methotrexate Resistance in the Protozoan Parasite Leishmania major

Trypanosomatid protozoans are pterin auxotrophs, a finding noted decades ago which heralded the discovery of key metabolic roles played by pteridines in eukaryotes. We have now identified the enzyme mediating unconjugated pteridine salvage in the human parasite Leishmania major, PTR1 (pteridine redu...

Full description

Saved in:
Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1994-11, Vol.91 (24), p.11442-11446
Main Authors: Bello, A R, Nare, B, Freedman, D, Hardy, L, Beverley, S M
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Biochemistry
Chromosomes
Cloning, Molecular
DNA
DNA Primers - chemistry
Drug Resistance
Enzymes
Genes, Protozoan
Genetics
Hypersensitivity
Leishmania major
Leishmania major - metabolism
Metabolism
Methotrexate - pharmacology
Molecular Sequence Data
Mutagenesis, Insertional
Oxidoreductases - genetics
Parasites
Pteridines
Pteridines - metabolism
Pterins
Transfection
Citations: Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Trypanosomatid protozoans are pterin auxotrophs, a finding noted decades ago which heralded the discovery of key metabolic roles played by pteridines in eukaryotes. We have now identified the enzyme mediating unconjugated pteridine salvage in the human parasite Leishmania major, PTR1 (pteridine reductase 1, formerly hmtxror ltdh). PTR1 is the gene in the amplified H region responsible for methotrexate (MTX) resistance, and belongs to a large family of oxidoreductases with diverse substrates and roles. We generated Leishmania lacking PTR1 by homologous gene targeting, and these ptr1-mutants required reduced biopterin (dihydro- or tetrahydrobiopterin) for growth. PTR1 purified from engineered Escherichia coli exhibited a MTX-sensitive, NADPH-dependent biopterin reductase activity. PTR1 showed good activity with folate and significant activity with dihydrofolate and dihydrobiopterin, but not with quinonoid dihydrobiopterin. PTR1 thus differs considerably from previously reported pteridine reductases of trypanosomatids and vertebrates. Pteridine reductase activity was diminished in ptr1-Leishmania and was elevated in transfected parasites bearing multiple copies of PTR1; correspondingly, ptr1-was MTX-hypersensitive whereas the multicopy transfectant was MTX-resistant. The concordance of the biochemical and genetic properties of PTR1 suggests that this is the primary enzyme mediating pteridine salvage. These findings suggest several possible mechanisms for PTR1-mediated MTX resistance and should aid in the design of rational chemotherapy.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.91.24.11442