The roles of pteridine reductase 1 and dihydrofolate reductase- thymidylate synthase in pteridine metabolism in the protozoan parasite Leishmania major

Bakela Nare, Larry W. Hardy, Stephen M. Beverley

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130 Scopus citations

Abstract

Trypanosomatid protozoans depend upon exogenous sources of pteridines (pterins or folates) for growth. A broad spectrum pteridine reductase (PTR1) was recently identified in Leishmania major, whose sequence places it in the short chain alcohol dehydrogenase protein family although its enzymatic activities resemble dihydrofolate reductases. The properties of PTR1 suggested a role in essential pteridine salvage as well as in antifolate resistance. To prove this, we have characterized further the properties and relative roles of PTR1 and dihydrofolate reductase-thymidylate synthase in Leishmania pteridine metabolism, using purified enzymes and knockout mutants. Recombinant L. major and Leishmania tarentolae, and native L. major PTR1s, were tetramers of 30-kDa subunits and showed similar catalytic properties with pterins and folates (pH dependence, substrate inhibition with H2pteridines). Unlike PTR1, dihydrofolate reductase-thymidylate synthase showed weak activity with folate and no activity with pterins. Correspondingly, studies of ptr1- and dhfr-ts- mutants implicated only PTR1 in the ability of L. major to grow on a wide array of pterins. PTR1 exhibited 2000-fold less sensitivity to inhibition by methotrexate than dihydrofolate reductase-thymidylate synthase, suggesting several mechanisms by which PTR1 may compromise antifolate inhibition in wild-type Leishmania and lines bearing PTR1 amplifications. We incorporate these results into a comprehensive model of pteridine metabolism and discuss its implications in chemotherapy of this important human pathogen.

Original languageEnglish
Pages (from-to)13883-13891
Number of pages9
JournalJournal of Biological Chemistry
Volume272
Issue number21
DOIs
StatePublished - May 23 1997
Externally publishedYes

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