Pteridine reductase 1 (PTR1, also known as HMTXR) is a cytosolic, NADPH-dependent short-chain dehydrogenase/reductase (SDR) that forms a homotetramer of 30-kDa subunits. Leishmania and other trypanosomatids cannot synthesize pterins de novo, and PTR1 is the main enzyme of unconjugated pterin salvage. It reduces imported biopterin in two successive steps, first to 7,8-dihydrobiopterin and then to 5,6,7,8-tetrahydrobiopterin (H4B). The first reduction follows the general SDR mechanism; the second resembles the dihydrofolate reductase (DHFR) mechanism. PTR1 has broad substrate specificity. It also reduces folate and dihydrofolate to tetrahydrofolate with the same stereochemistry as DHFR, and it has a lower, NADPH-dependent quinonoid dihydropteridine reductase activity. ptr1 null promastigotes need reduced biopterin to grow, and loss of PTR1 changes metacyclogenesis and sensitivity to oxidative stress. PTR1 is about 2000-fold less sensitive to methotrexate than DHFR-thymidylate synthase. It was first identified as the amplified H-region gene that confers methotrexate resistance, because it provides a route for folate reduction that bypasses antifolate inhibition, and it is a target for antitrypanosomatid drug design.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004155 6,7-dihydropteridine reductase activity | IDA PMID:21239486 Dissecting the metabolic roles of pteridine reductase 1 in T... | KEEP AS NON CORE | Summary: Ong et al. (2011) assayed recombinant LmPTR1 with an HPLC/spectrophotometric method and found an NADPH-dependent quinonoid dihydrobiopterin (qH2B) reductase activity, with a specificity constant similar to that for 7,8-dihydrobiopterin. Reason: The activity is directly demonstrated and supersedes the 1994 report of no activity with qH2B (PMID:7972081), so the annotation stands. In L. major, however, the dedicated NADH-dependent QDPR is about 20-fold more efficient with qH2B than LmPTR1. PTR1's quinonoid reductase activity is therefore a secondary, backup activity in Leishmania. It may matter more in T. brucei, which lacks a QDPR. The term is as specific as warranted but peripheral to the core salvage function. Supporting Evidence: PMID:21239486 we demonstrate that there is an NADPH-dependent QDPR activity associated with both TbPTR1 and LmPTR1. PMID:21239486 Specificity constants (k(cat)/K(m)) for LmPTR1 are similar with dihydrobiopterin (H(2)B) and quinonoid dihydrobiopterin (qH(2)B) as substrates and about 20-fold lower than LmQDPR with qH(2)B. In contrast, TbPTR1 shows a 10-fold higher k(cat)/K(m) for H(2)B over qH(2)B. |
| GO:0005829 cytosol | ISO GO_REF:0000024 | ACCEPT | Summary: Transferred from T. brucei PTR1 (Tb927.8.2210). In digitonin fractionation, TbPTR1 was recovered only in the cytosolic fraction. In L. donovani, GFP-tagged PTR1 is cytoplasmic. Reason: Orthologous data from T. brucei biochemical fractionation and Leishmania GFP tagging agree. PTR1 has no glycosomal targeting evidence, and the protein did not fractionate with glycosomes. Cytosol is the correct, specific compartment for this soluble salvage enzyme. Supporting Evidence: PMID:20545846 PTR1 was recovered exclusively in the cytosolic fraction along with the cytosolic enzyme marker, enolase. PMID:17275814 Fluorescence microscopy revealed cytoplasmic localization of GFP tagged protein extending to the flagellum in these stationary phase promastigotes, implying that degradation of the protein was not by the usual multivesicular tubule lysosome (MVT) pathway. |
| GO:0006729 tetrahydrobiopterin biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: The UniPathway mapping places PTR1 in the step "tetrahydrobiopterin from biopterin". PTR1 catalyzes the NADPH-dependent two-step reduction of biopterin to H4B, the only route to H4B in pterin-auxotrophic Leishmania. Reason: PTR1 itself catalyzes formation of H4B from salvaged biopterin, and ptr1 null parasites need reduced biopterin for growth. This is salvage, not the de novo GTP route, which trypanosomatids lack. The GO term covers any pathway that forms H4B, so it is correct and core. Supporting Evidence: PMID:11373620 PTR1 catalyzes the NADPH-dependent two-step reduction of oxidized pterins to the active tetrahydro-forms and reduces susceptibility to antifolates by alleviating dihydrofolate reductase (DHFR) inhibition. PMID:7972081 these ptr1- mutants required reduced biopterin (dihydro- or tetrahydrobiopterin) for growth. |
| GO:0016491 oxidoreductase activity | IBA GO_REF:0000033 | MODIFY | Summary: PAINT propagation of generic SDR oxidoreductase activity. Q01782 appears among its own donors because it has experimental annotations, as expected. Reason: The claim is correct; PTR1 is an NADPH-dependent SDR reductase. But the term is far less specific than the directly measured activities. Replace it with pteridine reductase activity (GO:0047040), which the gene already carries by IDA, and dihydrofolate reductase activity (GO:0004146), shown by kinetic and stereochemical studies. Both are is_a descendants of oxidoreductase activity. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002462421 · PTN002462421 SUPPORTS TRANSFER PTR1 is inside this SDR clade and generic oxidoreductase activity is conserved across it. Q01782 is among the donors because it has its own experimental evidence. The MODIFY only sharpens the term to the measured pteridine and dihydrofolate reductase activities; it does not dispute the transfer. Proposed replacements: pteridine reductase activity dihydrofolate reductase activity Supporting Evidence: PMID:7972081 PTR1 purified from engineered Escherichia coli exhibited a MTX-sensitive, NADPH-dependent biopterin reductase activity. PMID:9521731 With folate as a substrate, PTR1 catalyzes two rounds of reduction, yielding 5,6,7, 8-tetrahydrofolate and oxidizing 2 equiv of NADPH. |
| GO:0016491 oxidoreductase activity | ISM GO_REF:0000011 | MODIFY | Summary: An InterPro SDR family signature (IPR002198) maps to generic oxidoreductase activity. Reason: Same claim as the IBA row. It is correct but general, and the specific activities are experimentally established. Replace with GO:0047040 and GO:0004146. Proposed replacements: pteridine reductase activity dihydrofolate reductase activity Supporting Evidence: PMID:9521731 The kinetic mechanism and stereochemical course for the catalyzed reaction confirm PTR1's membership within the short chain dehydrogenase/reductase (SDR) family. |
| GO:0047040 pteridine reductase activity | IDA PMID:21239486 Dissecting the metabolic roles of pteridine reductase 1 in T... | ACCEPT | Summary: Ong et al. measured the kinetics of recombinant LmPTR1 with 7,8-dihydrobiopterin and NADPH at physiological pH and ionic strength. Reason: Pteridine reductase (EC 1.5.1.33) is the defining, core activity of PTR1. It is supported here and by the original biochemical and structural studies. Supporting Evidence: PMID:21239486 Leishmania parasites are pteridine auxotrophs that use an NADPH-dependent pteridine reductase 1 (PTR1) PMID:7972081 PTR1 purified from engineered Escherichia coli exhibited a MTX-sensitive, NADPH-dependent biopterin reductase activity. |
| GO:0047040 pteridine reductase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic mapping from the UniProt catalytic activity (RHEA:19509, EC 1.5.1.33). Reason: Consistent with the experimentally established core activity. Supporting Evidence: PMID:11373620 Pteridine reductase (PTR1) is a short-chain reductase (SDR) responsible for the salvage of pterins in parasitic trypanosomatids. |
| GO:0047040 pteridine reductase activity | ISO GO_REF:0000024 | ACCEPT | Summary: Transferred from the T. brucei ortholog PTR1 (Tb927.8.2210), which has the same activity. Reason: Redundant with the direct L. major evidence, but correct and consistent. Supporting Evidence: PMID:21239486 Leishmania parasites are pteridine auxotrophs that use an NADPH-dependent pteridine reductase 1 (PTR1) |
| GO:0004146 dihydrofolate reductase activity | IDA PMID:9521731 Leishmania major pteridine reductase 1 belongs to the short ... | NEW | Summary: PTR1 reduces folate to dihydrofolate and dihydrofolate to 5,6,7,8-tetrahydrofolate with NADPH. It transfers the pro-S hydride to C6 on the si face and gives the same THF stereoisomer as DHFR. Reason: GO:0004146 is defined by the reaction THF + NADP+ = DHF + NADPH, which PTR1 catalyzes directly as shown with purified enzyme. This folate-reducing activity underlies PTR1-mediated methotrexate resistance. GOA does not capture it for Q01782. PTR1 is the catalyst, so no participation concern arises. DHFR-TS is the main folate reductase, so this is a secondary, physiologically conditional activity. Supporting Evidence: PMID:9521731 With folate as a substrate, PTR1 catalyzes two rounds of reduction, yielding 5,6,7, 8-tetrahydrofolate and oxidizing 2 equiv of NADPH. PMID:9521731 PTR1 transfers the pro-S hydride of NADPH to carbon 6 on the si face of dihydrofolate, producing the same stereoisomer of THF as does dihydrofolate reductase. |
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Download this section (compressed HTML)Q: Given that LmPTR1 reduces quinonoid dihydrobiopterin with efficiency similar to 7,8-dihydrobiopterin, does PTR1 contribute measurably to H4B regeneration in L. major cells lacking QDPR, or is the dedicated QDPR sufficient?
Q: Which H4B-dependent reactions explain the requirement for reduced pterins in Leishmania, given that the parasite lacks NOS and the aromatic amino acid hydroxylases other than a non-essential PAH?
Q: What enzyme provides the residual pteridine reductase activity (PTR2) seen in ptr1 null Leishmania?
Experiment: Generate ptr1/qdpr double mutants in L. major supplemented with H4B and measure intracellular H4B/qH2B pools by HPLC, with and without complementation by a PTR1 variant selectively impaired for quinonoid substrates.
Hypothesis: PTR1 quinonoid reductase activity partially compensates for QDPR loss in vivo.
Experiment: Determine the localization of untagged endogenous PTR1 in L. major promastigotes and amastigotes by immunofluorescence and digitonin fractionation.
Hypothesis: Endogenous PTR1 is cytosolic and not glycosomal in both life stages.
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