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evolution
alignment analysis shows several substitutions between Schistosoma mansoni and human DHFR at both the folate- and NADP+-binding sites. The folate-binding site exhibits two differences at three residues (human DHFR numbering): E30D and N64F. The NADP+-binding site exhibits six substitutions at 11 residues: D21G,K54R, K55V, R76S, E77T and S118Y
evolution
dihydrofolate reductase (DHFR) and thymidylate synthase (TS) have undergone a fusion event generating a single polypeptide but conserving the two functions in trypanosomatids
evolution
dihydrofolate reductase (DHFR) and thymidylate synthase (TS) have undergone a fusion event generating a single polypeptide but conserving the two functions in trypanosomatids
evolution
dihydrofolate reductase (DHFR) and thymidylate synthase (TS) have undergone a fusion event generating a single polypeptide but conserving the two functions in trypanosomatids
evolution
identification of a second functional dihydrofolate reductase enzyme in humans, DHFRL1. RNA-mediated DHFR duplication events occur across the mammal tree. Dihydrofolate reductase activity is also a feature of the mitochondria in both rat and mouse but this is not due to a second enzyme. Humans have evolved the need for two separate enzymes, while laboratory rats and mice have just one. RNA-mediated DHFR duplicates in brown rat and mouse are likely to be processed pseudogenes
evolution
identification of a second functional dihydrofolate reductase enzyme in humans, DHFRL1. RNA-mediated DHFR duplication events occur across the mammal tree. Dihydrofolate reductase activity is also a feature of the mitochondria in both rat and mouse but this is not due to a second enzyme. Humans have evolved the need for two separate enzymes, while laboratory rats and mice have just one. RNA-mediated DHFR duplicates in brown rat and mouse are likely to be processed pseudogenes
evolution
identification of a second functional dihydrofolate reductase enzyme in humans, DHFRL1. RNA-mediated DHFR duplication events occur across the mammal tree. Dihydrofolate reductase activity is also a feature of the mitochondria in both rat and mouse but this is not due to a second enzyme. Humans have evolved the need for two separate enzymes, while laboratory rats and mice have just one. RNA-mediated DHFR duplicates in brown rat and mouse are likely to be processed pseudogenes
evolution
PTR1 is a NADPH-dependent enzyme belonging to the short-chain dehydrogenase/reductase (SDR) family
evolution
PTR1 is a short-chain dehydrogenase reductase family member. The trypanosomatid PTR1s are structurally very similar, sequence comparisons
evolution
PTR2 is a short-chain dehydrogenase reductase family member. In Trypanosoma cruzi, TcPTR1 and TcPTR2 are isoforms that show very high sequence homology but also display varied enzymatic activity. TcPTR1 in comparison to TcPTR2 shows higher activity with biopterin and folate than with H2F or H2B
evolution
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replacement of Asp27 in Escherichia coli DHFR by Glu27 in Moritella DHFR may be an adaptation for cold that fortuitously also enhances activity under pressure. The extra carbon of a glutamate increases flexibility of the Thr113-Res27 hydrogen bond while pressure increases the hydrogen bond strength and correlation of sheet F with helix B
evolution
the enzyme belong to the short-chain dehydrogenase/reductase (SDR) family of enzymes. Despite the overall low sequence identity among members of the SDR family (about 15-30%), a central catalytic YX3K motif is highly conserved, as is an N-terminal glycine motif (TGX3GXG), involved in cofactor binding and recognition. The pteridine reductases in the SDR family have an arginine in place of the glycine at position 6 in this motif (TGX3RXG)
evolution
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Tyr103 of Moritella yayanosii DHFR may be an adaptation for high pressure since Cys103 in helix F of Moritella profunda DHFR forms an intra-helix hydrogen bond with Ile99 while Tyr103 in helix F of MyDHFR forms a hydrogen bond with Leu78 in helix E. Tyr 103 may be an adaptation for preventing distortion of the adenosine binding domain at higher pressures that may only be advantageous for cold-adapted DHFRs
malfunction
inhibition of DHFR leads to the depletion of tetrahydrofolate and eventual cell death
malfunction
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inhibition of the enzyme activity leads to arrest of DNA synthesis and hence cell death
malfunction
inhibitor trimethoprim shows loss potency and NADPH synergy on binding S1 mutant DHFR. Mutation of residues Y98F/A43G in S1 mutant restores trimethoprim sensitivity and NADPH synergy
malfunction
enzyme knockdown results in the abnormal developments of zebrafish embryos in the early stages. Obvious malformations in heart and outflow tract are also observed in knockdown embryos. Enzyme knockdown causes reduced cell proliferation and increased apoptosis
malfunction
growth deficiency phenotypes (in un-supplemented M9 minimal medium containing thymidine) are direct consequences of the gene deletions
malfunction
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growth deficiency phenotypes (in un-supplemented M9 minimal medium containing thymidine) are direct consequences of the gene deletions
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metabolism
DHFR is a critical enzyme in the maintenance of reduced folate pools used in the biosynthetic pathways of purines, thymidylate, methionine, glycine, pantothenic acid, and N-formyl-methionyl tRNA
metabolism
dihydrofolate reductase (DHFR) is involved in the thymidylate and is essential for nucleotide metabolism
metabolism
enzyme DHFR is a pivotal enzyme in the thymidine and purine synthesis pathway
metabolism
the enzyme is involved in the folate recycling pathway
metabolism
enzyme displays a kinetic isotope effect KIE (kHLE/kHHE) close to unity above 0°C. The enzyme KIE is increased to 1.720 at -20°C, The coupling of protein motions to the chemical step may be minimized under optimal conditions but enhanced at non-physiological temperatures
metabolism
key enzymes involved in trypanosome folate metabolism are dihydrofolate reductase (DHFR) and pteridine reductase (PTR1)
metabolism
key enzymes involved in trypanosome folate metabolism are dihydrofolate reductase (DHFR) and pteridine reductase (PTR1)
metabolism
pteridine reductase 1 (PTR1) is required for tetrahydrobiopterin (H4BPt) synthesis
metabolism
study of protein dynamics, using a pump-probe method that employs pulsed-laser photothermal heating of a gold nanoparticle (AuNP) to directly excite a local region of the protein structure and transient absorbance to probe the effect on enzyme activity. Activity is accelerated by pulsed-laser excitation when the AuNP is attached close to a network of coupled motions in DHFR. No rate acceleration is observed when the AuNP is attached away from the network with pulsed excitation, or for any attachment site with continuous wave excitation
metabolism
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the rate of hydride transfer is the rate-determining step at the basic pH region, and is independent of salt concentration. The amplitude of the DHF-binding reaction increases and the tetrahydrofolate releasing rate decreases with increasing NaCl concentration
metabolism
traditional antifolates, such as methotrexate (MTX) inhibiting DHFR, are poorly effective towards trypanosome parasites because of the metabolic bypass provided by PTR1 also catalyzing folate reduction in addition to the conversion of biopterin to 7,8-dihydrobiopterin (DHB) and subsequently to 5,6,7,8-tetrahydrobiopterin (THB)
metabolism
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the enzyme is involved in the folate recycling pathway
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metabolism
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the rate of hydride transfer is the rate-determining step at the basic pH region, and is independent of salt concentration. The amplitude of the DHF-binding reaction increases and the tetrahydrofolate releasing rate decreases with increasing NaCl concentration
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physiological function
DHFR is essential for the survival and pathogenesis of anthrax. DHFR is required for de novo DNA synthesis and amino acid synthesis
physiological function
DHFR is the enzyme responsible for the NADPH-dependent reduction of 5,6-dihydrofolate to 5,6,7,8-tetrahydrofolate, an essential cofactor in the synthesis of purines, thymidylate, methionine, and other key metabolites. DHFR is a critical enzyme in the maintenance of reduced folate pools used in the biosynthetic pathways of purines, thymidylate, methionine, glycine, pantothenic acid, and N-formyl-methionyl tRNA
physiological function
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dihydrofolate reductase is an enzyme with a pivotal role in the synthesis of intracellular tetrahydrofolic acid, which is essential in the synthesis of purines, some amino acids, and thymidine. DHFR is the sole source of tetrahydrofolic acid
physiological function
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dihydrofolate reductase is an enzyme with a pivotal role in the synthesis of intracellular tetrahydrofolic acid, which is essential in the synthesis of purines, some amino acids, and thymidine. DHFR is the sole source of tetrahydrofolic acid
physiological function
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dihydrofolate reductase is an enzyme with a pivotal role in the synthesis of intracellular tetrahydrofolic acid, which is essential in the synthesis of purines, some amino acids, and thymidine. DHFR is the sole source of tetrahydrofolic acid
physiological function
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tetrahydrofolate is a precursor of cofactors necessary for the synthesis of thymidylate, purine nucleotides,methionine, serine, and glycine required for DNA, RNA, and protein synthesis
physiological function
dihydrofolate reductase is required for the development of heart and outflow tract in zebrafish. Enzyme overexpression promotes cell proliferation and inhibited apoptosis
physiological function
DHFR-TS is essential for cell survival of Trypanosoma brucei
physiological function
dihydrofolate reductase (DHFR) is an enzyme from the folate one-carbon metabolism pathway that plays a role in drug resistance and in reducing the synthetic supplement folic acid and 7,8-dihydrofolate to the active form, tetrahydrofolate
physiological function
dihydrofolate reductase (DHFR) is an enzyme from the folate one-carbon metabolism pathway that plays a role in drug resistance and in reducing the synthetic supplement folic acid and 7,8-dihydrofolate to the active form, tetrahydrofolate
physiological function
dihydrofolate reductase (DHFR) is an enzyme from the folate one-carbon metabolism pathway that plays a role in drug resistance and in reducing the synthetic supplement folic acid and 7,8-dihydrofolate to the active form, tetrahydrofolate
physiological function
dihydrofolate reductase is an essential enzyme in the tetrahydrofolate pathway which catalyzes the NADPH-dependent reduction of 7,8-dihydrofolate (H2F) to the 5,6,7,8-tetrahydrofolate needed to maintain intracellular pools of tetrahydrofolate and its derivatives. These are essential cofactors in the biosynthesis of purines, pyrimidines and several amino acids
physiological function
dihydrofolate reductase is an essential enzyme in the tetrahydrofolate pathway which catalyzes the NADPH-dependent reduction of 7,8-dihydrofolate (H2F) to the 5,6,7,8-tetrahydrofolate needed to maintain intracellular pools of tetrahydrofolate and its derivatives. These are essential cofactors in the biosynthesis of purines, pyrimidines and several amino acids
physiological function
dihydrofolate reductase is an essential enzyme in the tetrahydrofolate pathway which catalyzes the NADPH-dependent reduction of 7,8-dihydrofolate (H2F) to the 5,6,7,8-tetrahydrofolate needed to maintain intracellular pools of tetrahydrofolate and its derivatives. These are essential cofactors in the biosynthesis of purines, pyrimidines and several amino acids
physiological function
several parasitic protozoa, including Toxoplasma gondii, contain a unique bifunctional thymidylate synthase-dihydrofolate reductase (TS-DHFR) having the catalytic activities contained on a single polypeptide chain in contrast to the human enzyme. Three-dimensional structures of Toxoplasma gondii enzyme TS-DHFR and of a loop truncated TS-DHFR enzyme, removing several flexible surface loops in the DHFR domain, shows that the TS-DHFR homodimer includes a junctional region containing a linked crossover helix between the DHFR domains of the two adjacent monomers, a long linker connecting the TS and DHFR domains, and a DHFR domain that is positively charged. The crystal structure suggests that the positively charged DHFR domain governs this electrostatically mediated movement of dihydrofolate, preventing release from the enzyme. Importance of this region not only in DHFR catalysis but also in modulating the distal TS activity suggests a role for TS-DHFR interdomain interactions
physiological function
the human thymidylate synthase and dihydrofolate reductase catalyse two consecutive reactions in the folate metabolism pathway and are very likely to bind in the same multi-enzyme complex in vivo, substrate channeling occurs between the human thymidylate synthase and dihydrofolate reductase, analysis by protein-protein docking, electrostatics calculations, and Brownian dynamics, overview. The non-covalently non-covalently bound human thymidylate synthase and dihydrofolate reductase are capable of substrate channeling and the formation of the surface electrostatic highway. The substrate channeling efficiency between the two can be reasonably high and comparable to that of the bifunctional protozoan enzyme
physiological function
DHFR is essential for growth in vitro. Nutritional supplements of most forms of folate are not sufficient to restore growth when DHFR expression is suppressed or when its activity is directly inhibited by methotrexate. A strain with doxycycline-repressible expression of DHFR is rendered avirulent in a mouse model of disseminated candidiasis upon doxycycline treatment
physiological function
enzyme is able to complement a Saccharomyces cerevisiae DHFR mutant strain
physiological function
for mechanism, dynamics are crucial for solvent entry and protonation of substrate. The mechanism invokes the release of a sole proton from a hydronium (H3O+) ion, its pathway through a narrow channel that sterically hinders the passage of water, and the ultimate protonation of DHF at the N5 atom. DOD47 is the catalytic water that promotes protonation of the N5 atom in DHF. The deuteron, modeled into the nuclear difference density peak in the active site, possibly forms a low-barrier hydrogen bond with the oxygen atom of DOD47 and is positioned between the Met20 side chain and the DOD47 so as to define a pathway that could lead to protonation of N5 by the deuteron
physiological function
in addition to folate reduction, PTR1 (EC 1.5.1.33) also catalyzes the conversion of biopterin to 7,8-dihydrobiopterin (DHB) and subsequently to 5,6,7,8-tetrahydrobiopterin (THB). Under dihydrofolate reductase (DHFR) inhibition, PTR1 is upregulated providing reduced folates necessary for parasite survival
physiological function
pteridine reductase 1 (PTR1) has the ability to catalyze the NADPH-dependent two-stage reduction of biopterins to their 7,8-dihydro and 5,6,7,8-tetrahydro forms as well as folates to their H2F and H4F forms. PTR1 is a trypanosomatid multifunctional enzyme that provides a mechanism for escape of dihydrofolate reductase (DHFR, EC 1.5.1.3) inhibition. This is because PTR1 can reduce pterins and folates. Trypanosomes require folates and pterins for survival and are unable to synthesize them de novo
physiological function
pteridine reductase 1 (PTR1, EC 1.5.1.33) has the ability to catalyze the NADPH-dependent two-stage reduction of biopterins to their 7,8-dihydro and 5,6,7,8-tetrahydro forms as well as folates to their H2F and H4F forms. PTR1 is a trypanosomatid multifunctional enzyme that provides a mechanism for escape of dihydrofolate reductase (DHFR) inhibition. This is because PTR1 can reduce pterins and folates. Trypanosomes require folates and pterins for survival and are unable to synthesize them de novo. In Trypanosoma cruzi, TcPTR1 and TcPTR2 are isoforms that show very high sequence homology but also display varied enzymatic activity. TcPTR1 in comparison to TcPTR2 shows higher activity with biopterin and folate than with dihydrofolate or dihydrobiopterin
physiological function
PTR1 produces tetrahydrobiopterin (H4BPt) from dihydrobiopterin (H2BPt) or from fully oxidized biopterin (BPt) scavenged from host cells
physiological function
the bifunctional dihydrofolate reductase-thymidylate synthase, DHFR-TS, enzyme includes dthe ihydrofolate reductase, EC 1.5.1.3, and the thymidylate synthase, EC 2.1.1.45. Under dihydrofolate reductase (DHFR) inhibition, the PTR1 (EC 1.5.1.33) gene is upregulated, providing reduced folates necessary for parasite survival
physiological function
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DHFR-TS is essential for cell survival of Trypanosoma brucei
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additional information
analysis of the orientation of residues Y100, D27, M20, and the ligands in the active site of ecDHFR, PDB ID 1rx2
additional information
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analysis of the orientation of residues Y100, D27, M20, and the ligands in the active site of ecDHFR, PDB ID 1rx2
additional information
DHFR domain structure analysis, the domain consists of 252 residues from the N-terminus to the start of the junctional region, overview
additional information
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DHFR domain structure analysis, the domain consists of 252 residues from the N-terminus to the start of the junctional region, overview
additional information
homology model of enzyme pjDHFR in ternary complex with NADPH and inhibitor 2,4-diamino-6-[(2',5'-dichloro anilino)methyl]pyrido[2,3-d]pyrimidine
additional information
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homology model of enzyme pjDHFR in ternary complex with NADPH and inhibitor 2,4-diamino-6-[(2',5'-dichloro anilino)methyl]pyrido[2,3-d]pyrimidine
additional information
in vitro, one human thymidylate synthase dimer binds to up to six human dihydrofolate reductase monomers, protein-protein docking process, overview. Existence of bound-state conformations of the human DHFR and TS proteins where a continuous positive surface potential region, connecting the TS and DHFR active sites, is formed
additional information
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in vitro, one human thymidylate synthase dimer binds to up to six human dihydrofolate reductase monomers, protein-protein docking process, overview. Existence of bound-state conformations of the human DHFR and TS proteins where a continuous positive surface potential region, connecting the TS and DHFR active sites, is formed
additional information
role for water in stabilizing the DHFR substrate pocket and of Cys52, overview
additional information
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role for water in stabilizing the DHFR substrate pocket and of Cys52, overview
additional information
the DHFR enzyme structure exhibits the canonical DHFR fold
additional information
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the DHFR enzyme structure exhibits the canonical DHFR fold
additional information
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the predicted specificity determining positions in BmDHFR for ligand binding are Val5, Gly12, Met27, Asn44, Ala45, Lys51, Phe60, Val84, Phe89 Leu95, Leu96, Gly133, Ala137, Val139, Phe140, Phe141 and Glu169
additional information
enzyme structure modelling
additional information
isozyme TcPTR1 has no crystal structure, so for this study a structure is calculated using homology modeling with TcPTR2 as a template
additional information
pterin and folate substrates, along with inhibitors, interact with PTR1 complexes quite similarly, often via binding in a Pi-sandwich between the NADPH nicotinamide ring and residue Phe97. The NADPH cofactor is known to be essential in creating both the substrate binding site as well as the catalytic center. Arg14, Ser95, Phe97, Asp161, and Tyr174 are important residues that interact with the folate and pterin substrates
additional information
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role for water in stabilizing the DHFR substrate pocket and of Cys52, overview
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