| 2000 |
TRX1 (PfTrx, 13 kDa) was cloned, expressed, and biochemically characterized. It has a midpoint redox potential of -270 mV and is reduced by both P. falciparum TrxR (Km = 10.4 µM, kcat = 3100 min⁻¹) and human TrxR (Km = 2 µM, kcat = 3300 min⁻¹). Reduced TRX1 reduces GSSG with a rate constant k2 = 0.039 µM⁻¹ min⁻¹, approximately 5-fold faster than thioredoxins from human, Drosophila, or E. coli, and also reduces S-nitrosoglutathione. |
Recombinant protein expression, steady-state enzyme kinetics, redox potential measurement |
The Journal of biological chemistry |
High |
11013257
|
| 2001 |
TRX1 active-site cysteines (Cys30 and Cys33) are both essential for catalytic activity as shown by mutational analyses. A third cysteine (Cys43) outside the active site is partially responsible for dimer formation; mutating Cys43 to Ala reduces dimerization. All three cysteines are accessible to DTNB modification, with Cys33 more reactive than Cys30. Steady-state kinetics with PfTrxR give Km values of 0.8–2.1 µM for TRX1, ~250-fold lower than for E. coli thioredoxin. |
Site-directed mutagenesis, DTNB reactivity assay, steady-state kinetics with recombinant PfTrxR |
Molecular and biochemical parasitology |
High |
11223129
|
| 2006 |
The mitochondrial peroxiredoxin of P. falciparum strongly prefers the mitochondrial thioredoxin (PfTrx2) over the cytosolic TRX1 as a reducing partner (Km 11.6 µM vs. 130.4 µM for TRX1). Substitution of residues Asn62/Tyr63 and Phe139/Ala140 in the peroxiredoxin reduced the Km for TRX1 but had no effect on Km for PfTrx2, indicating these residues mediate substrate discrimination. |
Recombinant protein expression, steady-state kinetics, site-directed mutagenesis of the peroxiredoxin |
Molecular microbiology |
High |
16879648
|
| 2009 |
Human peroxiredoxin 2 (hPrx-2), imported into the P. falciparum cytosol, accepts cytosolic TRX1 as a reducing substrate in enzyme kinetic analyses, accounting for roughly 50% of thioredoxin peroxidase activity in parasite extracts. |
Enzyme kinetic analysis with recombinant proteins, confocal microscopy, immunogold electron microscopy |
Proceedings of the National Academy of Sciences of the United States of America |
High |
19666612
|
| 2009 |
Using an active-site mutant of TRX1 lacking the resolving cysteine (to trap substrates via mixed disulfide), 17 putative redox-regulated target proteins of TRX1 were identified by affinity chromatography and mass spectrometry, including proteins in protein biosynthesis, energy metabolism, and signal transduction. TRX1 binding to S-adenosyl-L-homocysteine hydrolase and ornithine aminotransferase (OAT) was verified by surface plasmon resonance (BIAcore), and TRX1 was shown to enhance OAT enzymatic activity in functional assays. |
Active-site mutant affinity chromatography, mass spectrometry, BIAcore surface plasmon resonance, enzymatic activity assay |
PLoS pathogens |
High |
19360125
|
| 2010 |
TRX1 localizes to the cytoplasm of P. falciparum blood-stage parasites. Using alternative translation initiation, thioredoxin reductase (TrxR) achieves dual cytosolic and mitochondrial localization, providing TRX1's electron donor in the cytosol. |
Systematic subcellular localization study using GFP fusions and confocal microscopy |
PLoS pathogens |
Medium |
21203490
|
| 2010 |
TRX1 activates P. falciparum OAT activity approximately 10-fold. Crystal structure of OAT and site-directed mutagenesis identified Cys154 and Cys163 (within the substrate-binding loop, absent in human OAT) as the residues that mediate interaction with TRX1; the Cys154→Ser mutant shows strongly reduced OAT activity due to impaired ornithine binding. |
Crystal structure of OAT (2.3 Å), site-directed mutagenesis of OAT cysteines, enzymatic activity assays with recombinant TRX1 |
Journal of molecular biology |
High |
20673832
|
| 2010 |
Affinity chromatography with a resolving-cysteine mutant of TRX1 (C33S) captured 20 target proteins from P. falciparum extracts, including four glycolytic enzymes (hexokinase, GAPDH, phosphoglycerate mutase, LDH). TRX1 enhanced the enzymatic activity of PfGAPDH and PfLDH in functional assays. |
Cysteine-mutant affinity chromatography, LC-MS/MS, enzymatic activity assay |
Parasitology international |
Medium |
20307687
|
| 2011 |
TRX1 (thioredoxin 1) efficiently catalyzes protein deglutathionylation in P. falciparum, demonstrated by enzymatic deglutathionylation assays. TRX1 was itself identified as a target of S-glutathionylation in the parasite glutathionylome. |
Enzymatic deglutathionylation assay, affinity purification with biotin-maleimide, proteomic analysis by mass spectrometry |
Antioxidants & redox signaling |
Medium |
21595565
|
| 2013 |
Crystal structure of the P. falciparum TrxR–TRX1 complex was solved, revealing that complex formation induces rearrangements of the TrxR flexible C-terminal arm and an insertion loop. Amino acid residues at the TrxR–TRX1 interface were functionally analyzed by mutagenesis. The structure shows how TRX1 is bound and reduced by PfTrxR, with key differences from the human TrxR–thioredoxin interface. |
X-ray crystallography (crystal structure of TrxR–TRX1 complex), active-site mutagenesis, cell biological inhibitor experiments |
Journal of molecular biology |
High |
23845423
|
| 2014 |
TRX1 (PfTrx1) can be S-nitrosylated at its non-active-site cysteine (Cys43). Mechanistic studies demonstrated that TRX1 possesses both denitrosylating activity (mediated by active-site cysteines Cys30/Cys33) and transnitrosylating activity (mediated by Cys43). |
Biotin-switch assay, mass spectrometry, mechanistic mutagenesis/functional assays |
Antioxidants & redox signaling |
Medium |
24256207
|
| 2024 |
Solution structures of TRX1 in both reduced and oxidized forms were determined by high-resolution NMR. The protein adopts an α/β-fold with a mixed five-stranded β-sheet sandwiched between 4 helices (β1α1β2α2β3α3β4β5α4 topology). Oxidation of the CGPC active-site motif (Cys30–Cys33 disulfide) induces significant structural changes in residues Phe25–Ile36, Thr70–Thr74, and Leu88–Asn91. High-pressure NMR identifies at least three coexisting conformational sub-states in both redox forms. |
High-resolution NMR (solution structure determination of ¹³C/¹⁵N-labeled protein), high-pressure NMR spectroscopy |
PloS one |
High |
38635664
|