A rapid assay for dihydropteroate synthase activity suitable for identification of inhibitors

Fernley, Ross T.; Iliades, Peter; Macreadie, Ian

doi:10.1016/j.ab.2006.10.036

Cited by 7 publications

(5 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…There are multiple targets within the folate synthesis pathway, including dihydrofolate reductase (DHFR), a precursor shared between eukaryotic and prokaryotic organisms, as well as dihydropteroate synthase (DHPS), an enzyme unique to prokaryotic organisms [78]. Presently known inhibitors of these two targets interact with enzymatic neighbours in the folate pathway and include sulphonamides [78].…”

Section: The Folate Biosynthesis Pathway Represents a Mechanism Of Acmentioning

confidence: 99%

See 1 more Smart Citation

Bacteriocins and their position in the next wave of conventional antibiotics

Cavera

Arthur

Kashtanov

et al. 2015

International Journal of Antimicrobial Agents

161

View full text Add to dashboard Cite

Section: The Folate Biosynthesis Pathway Represents a Mechanism Of Acmentioning

confidence: 99%

“…Presently known inhibitors of these two targets interact with enzymatic neighbours in the folate pathway and include sulphonamides [78].…”

Section: The Folate Biosynthesis Pathway Represents a Mechanism Of Acmentioning

confidence: 99%

Bacteriocins and their position in the next wave of conventional antibiotics

Cavera

Arthur

Kashtanov

et al. 2015

International Journal of Antimicrobial Agents

161

View full text Add to dashboard Cite

“…Although, the assay can detect alternative substrates (though in the absence of pABA), it requires the separation of the pterin-sulfa adduct by chromatography to test for the possibility of DHFR inhibition. The assay described by Fernley et al [22], has a major advantage of using one coupling enzyme (DHFR); however, with the price of introduction of some ambiguities regarding the activity of DHFR in the presence of DHPS products.…”

Section: Discussionmentioning

confidence: 99%

“…An alternative method patented by Stachyra and Biton [21] could detect adduct formation because it detects PP i production, but it too is discontinuous and not suited for high throughput. More recently, an assay of DHPS was described by Fernley et al [22], which uses spectrophotometric detection making it amenable to high-throughput screening, but it detects the formation of dihydropteroate using the NADP + -dependent dihydrofolate reductase (DHFR) as coupling enzyme. Similar to the radiologic method with labeled pABA, it cannot directly account for assay interference by alternative substrates and adduct formation.…”

Section: Discussionmentioning

confidence: 99%

Examination of intrinsic sulfonamide resistance in Bacillus anthracis: A novel assay for dihydropteroate synthase

Valderas

Andi

Barrow

et al. 2008

Biochimica et Biophysica Acta (BBA) - General Subjects

View full text Add to dashboard Cite

Dihydropteroate synthase (DHPS) catalyzes the formation of dihydropteroate and Mgpyrophosphate from 6-hydroxymethyl-7,8-dihydropterin diphosphate and para-aminobenzoic acid. The Bacillus anthracis DHPS is intrinsically resistant to sulfonamides. However, using a radioassay that monitors the dihydropteroate product, the enzyme was inhibited by the same sulfonamides. A continuous spectrophotometric assay for measuring the enzymatic activity of DHPS was developed and used to examine the effects of sulfonamides on the enzyme. The new assay couples the production of MgPP i to the pyrophosphate-dependent phosphofructokinase/aldolase/triose isomerase/α-glycerophosphate dehydrogenase reactions and monitoring the disappearance of NADH at 340 nm. The coupled enzyme assay demonstrates that resistance of the B. anthracis DHPS results in part from the use of the sulfonamides as alternative substrates, resulting in the formation of sulfonamide-pterin adducts, and not necessarily due to an inability to bind them.

show abstract

“…Folates play a critical role in both DNA and RNA synthesis in cells [1][2][3][4][5][6]. Insu cient dietary intake of folates or intestinal absorption, or defective folate metabolism disrupt purine and pyrimidine nucleotide synthesis, especially for rapidly proliferating tissues such as hematopoietic and immune systems [7,8].…”

Section: Introductionmentioning

confidence: 99%

Immunodeficiency associated with a novel functionally defective variant of SLC19A1 benefits from folinic acid treatment

et al. 2022

View full text Add to dashboard Cite

Insu cient dietary folate intake, hereditary malabsorption, or defects in folate metabolism may lead to combined immunode ciency (CID). Although loss of function mutations in the major intestinal folate transporter PCFT/SLC46A1 was shown to be associated with CID, the evidence for pathogenic variants of RFC/SLC19A1 resulting in immunode ciency was lacking. We report two cousins carrying a homozygous pathogenic variant c.1042G>A, resulting in p.G348R substitution who showed symptoms of immunode ciency associated with defects of folate metabolism. SLC19A1 expression by peripheral blood mononuclear cells (PBMC) was quanti ed by real-time qPCR and immunostaining. T cell proliferation, methotrexate resistance, NK cell cytotoxicity, Treg cells and cytokine production by T cells were examined by ow cytometric assays. Patients were treated with and bene ted from folinic acid (FA). Studies revealed normal NK cell cytotoxicity, Treg cell counts, and naive-memory T cell percentages. Although SLC19A1 mRNA and protein expression were unaltered, remarkably, mitogen induced-T cell proliferation was signi cantly reduced at sub-and supraoptimal folic acid concentrations. In addition, patients' PBMCs were resistant to methotrexate-induced apoptosis supporting a functionally defective SLC19A1.This study presents the second pathogenic SLC19A1 variant in the literature, providing the rst experimental evidence that loss of function mutations in SLC19A1 may present with symptoms of immunode ciency.

show abstract

A rapid assay for dihydropteroate synthase activity suitable for identification of inhibitors

Cited by 7 publications

References 24 publications

Bacteriocins and their position in the next wave of conventional antibiotics

Bacteriocins and their position in the next wave of conventional antibiotics

Examination of intrinsic sulfonamide resistance in Bacillus anthracis: A novel assay for dihydropteroate synthase

Immunodeficiency associated with a novel functionally defective variant of SLC19A1 benefits from folinic acid treatment

Contact Info

Product

Resources

About