An assay for glucosamine-6-phosphate synthase using a yeast glucosamine-6-phosphate N-acetyltransferase 1 (GNA1) as coupling enzyme was developed. GNA1 transfers the acetyl moiety from acetyl-coenzyme A (CoA) to glucosamine-6-phosphate, releasing coenzyme A. The assay measures the production of glucosamine-6-phosphate by either following the consumption of acetyl-CoA spectrophotometrically at 230nm or quantifying the free thiol with 5,5'-dithio-bis(2-nitrobenzoic acid) (Ellman's reagent) in a discontinuous manner. This method is simple to perform and can be adapted to a 96-well microtiter plate format, which will facilitate high-throughput inhibitor screening and mechanistic studies using purified GlmS.

• PMID: 17880906
• DOI full text
• PubMed
• PubTator

Reference Type

Journal Article | Research Support, Non-U.S. Gov't

Authors

Li Y, Lopez P, Durand P, Ouazzani J, Badet B, Badet-Denisot MA

Primary Lit For

Additional Lit For

GNA1

Review For