title: Identification of uric acid as the redox molecule secreted by the yeast Arxula adeninivorans.
abstract: The yeast Arxula adeninivorans has been previously shown to secrete a large amount of an electro-active molecule. The molecule was produced by cells that had been cultivated in a rich medium, harvested, washed and then suspended in phosphate-buffered saline (PBS). The molecule was easily detectable after 60 min of incubation in PBS, and the cells continued to produce the molecule in these conditions for up to 3 days. The peak anodic potential of the oxidation peak was 0.42 V, and it was shown to be a solution species rather than a cell-attached species. We have optimised the production of the molecule, identified it by high-pressure liquid chromatography (HPLC) fractionation and high-resolution mass spectrometric analysis and determined the pathway involved in its synthesis. It has a mass/charge ratio that corresponds to uric acid, and this identification was supported by comparing UV spectra and cyclic voltammograms of the samples to those of uric acid. An A. adeninivorans xanthine oxidase gene disruption mutant failed to produce uric acid, which added further validity to this identification. It also demonstrated that the purine catabolism pathway is involved in its production. A transgenic A. adeninivorans strain with a switchable urate oxidase gene (AUOX) accumulated uric acid when the gene was switched off but did not when the gene was switched on. Cultivation of cells on amino acid and purine-free minimal media with an inorganic nitrogen source suggests that the cells synthesise purines from inorganic nitrogen and proceed to degrade them via the normal purine degradation pathway.
authors.affiliation: School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Williams
authors.forename: Anke
authors.initials: A
authors.lastname: Trautwein-Schult
authors.forename: Dagmara
authors.initials: D
authors.lastname: Jankowska
authors.forename: Gotthard
authors.initials: G
authors.lastname: Kunze
authors.forename: Marie A
authors.initials: MA
authors.lastname: Squire
authors.forename: Keith
authors.initials: K
authors.lastname: Baronian
mesh_terms: D002851:Chromatography, High Pressure Liquid; D003470:Culture Media / Q000737:chemistry; D017353:Gene Deletion; D015870:Gene Expression; D013058:Mass Spectrometry; D053858:Metabolic Networks and Pathways / Q000235:genetics; D009584:Nitrogen / Q000378:metabolism; D010084:Oxidation-Reduction; D011687:Purines / Q000378:metabolism; D004718:Saccharomycetales / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D014503:Urate Oxidase / Q000235:genetics / Q000378:metabolism; D014527:Uric Acid / Q000378:metabolism*; D014969:Xanthine Oxidase / Q000235:genetics / Q000378:metabolism
chemical_list: D003470:Culture Media; D011687:Purines; D014527:Uric Acid; D014969:Xanthine Oxidase; D014503:Urate Oxidase; D009584:Nitrogen