| 1985 |
GCH1 (GTP cyclohydrolase I) was isolated and partially purified (~200-fold) from P. falciparum erythrocytic-stage parasites. The enzyme catalyzes conversion of GTP to D-erythro-dihydroneopterin triphosphate (releasing carbon-8 of GTP). It has a native molecular weight of ~300,000 Da, optimal activity at pH 8.0 and 42°C, Km for GTP of 54.6 µM, and is Mg2+-independent. ATP is a competitive inhibitor (Ki = 600 µM); Mn2+, Cu2+, and Hg2+ are inhibitory. GCH1 activity in P. knowlesi is stage-dependent. |
Enzyme purification by size-exclusion HPLC, biochemical assay (fluorometric/spectrophotometric measurement of product), competitive inhibitor kinetics, product identification |
Molecular and biochemical parasitology |
High |
3908934
|
| 1989 |
P. falciparum synthesizes pteroylpolyglutamate de novo from GTP (the substrate of GCH1), p-aminobenzoate, and L-glutamate, establishing GCH1 as the entry point of an active de novo folate biosynthesis pathway in blood-stage parasites. The antifolates pyrimethamine and sulfadoxine at therapeutic concentrations inhibit incorporation of radiolabelled GTP into pteroylpentaglutamate, consistent with pathway inhibition upstream of or at GCH1. |
Metabolic radiolabelling with [14C]-GTP, HPLC product identification, drug inhibition assay |
Molecular and biochemical parasitology |
Medium |
2643036
|
| 2001 |
The gene encoding GCH1 was cloned, assigned unambiguously to P. falciparum chromosome 12, and shown to be expressed as a transcript in blood-stage parasites. The predicted protein sequence displays differences from the human host homologue that may be of functional significance. |
Gene cloning, chromosome assignment, Northern blot / RT-PCR transcript detection |
Parasitology |
Medium |
11197757
|
| 2002 |
GCH1 transcript levels in blood-stage P. falciparum change across the erythrocytic cell cycle and respond to folate pathway-specific stress (removal of metabolites, or challenge by folate enzyme inhibitors), but show no transcriptional feedback from downstream pathway inhibition. Expression is not exclusively constitutive/housekeeping. |
Quantitative RT-PCR (threshold detection) across the erythrocytic cell cycle in two cloned lines under normal and stress conditions |
Molecular microbiology |
Medium |
12366841
|
| 2011 |
Soluble, functional P. falciparum GCH1 (PfGCHI) could be heterologously expressed in bacteria when co-expressed with the P. falciparum chaperone PfHsp70. Sequential expression of PfHsp70 followed by PfGCHI increased yield of soluble, active enzyme. Purified His-tagged PfGCHI had a specific activity of 5.93 nmol/h/mg as measured by HPLC with fluorometric detection. |
Heterologous co-expression in E. coli, Ni-affinity chromatography purification, HPLC/fluorometric enzymatic activity assay |
Protein expression and purification |
Medium |
21262365
|
| 2013 |
Experimental manipulation of GCH1 copy number (CN) in P. falciparum parasite lines demonstrated that increased GCH1 CN directly alters pyrimethamine resistance in most parasite lines, providing direct experimental evidence that GCH1 expression level modulates antifolate drug susceptibility. Very high levels of GCH1 overexpression had a detrimental effect in lines with already high endogenous GCH1 expression, implicating GCH1 in maintaining metabolic flux balance in the folate pathway. |
Experimental CN manipulation by transfection, drug susceptibility assay (IC50 measurement), comparison across multiple parasite lines |
Molecular microbiology |
High |
23347134
|
| 2014 |
GCH1 amplification confers low-level pyrimethamine resistance and reduces the fitness cost of acquiring drug-resistant DHFR mutations downstream in the folate pathway. Epistasis analysis showed that GCH1 amplification compensates for compromised fitness resulting from stepwise DHFR mutations, expanding accessible evolutionary trajectories toward high-level antifolate resistance. |
Evolutionary trajectory analysis, parasite line comparison, growth fitness assays, epistasis analysis of GCH1 CN versus dhfr mutation background |
Molecular biology and evolution |
Medium |
24739308
|
| 2014 |
P. falciparum GCH1 was cloned, expressed in bacteria, and biochemically characterized. Enzymatic activity was measured by a fluorescence-based neopterin assay and validated with the GTP analogue 8-oxo-GTP (confirmed inhibitor). Domain truncation experiments using genetic complementation of ΔfolE E. coli showed that the conserved core domain of GCH1 is required for enzymatic activity. Homology modelling placed P. falciparum GCH1 in the Tunnelling-fold structural family with conserved catalytic residues at the active site. |
Recombinant protein expression, fluorescence-based enzymatic assay, 8-oxo-GTP inhibitor validation, domain truncation by genetic complementation in ΔfolE E. coli, homology modelling |
Malaria journal |
High |
24745605
|
| 2021 |
Normal mode analysis (anisotropic network model) of the P. falciparum GCH1 structure revealed that its tunnel-gating mechanism is regulated by a global shear motion and outward expansion of the central five-helix bundle. Hotspot residues within sites of structural significance for allosteric modulation were identified, providing a mechanistic basis for potential allosteric drug targeting. |
Computational normal mode analysis (anisotropic network model) of GCH1 crystal structure, alone and in complex with regulatory partner |
Journal of chemical information and modeling |
Low |
34450011
|
| 2025 |
Long-read sequencing of P. falciparum lines selected with a DHODH inhibitor revealed tandem GCH1 amplicons (up to 9 copies vs. 3 in parental lines). Expansions occurred in precise 2-unit steps with conserved AT-rich sequences at amplicon boundaries, consistent with the Plasmodium CNV formation model. A positive association was observed between GCH1 and DHODH copy number across independent selection experiments, suggesting GCH1 amplification may support DHODH locus amplification in the context of DHODH-inhibitor selection, linking folate and pyrimidine biosynthesis pathways. |
Long-read (Oxford Nanopore) sequencing, single-read visualization, copy number quantification, analysis of prior DHODH inhibitor selection experiments |
BMC genomics |
Medium |
40676505
|