Production of Arabidopsis thaliana UDP-Sugar Pyrophosphorylase by Pichia pastoris and Its Application in Efficient UDP-Glucose and UDP-Glucuronic Acid Synthesis

Resumo

Sugar nucleotides, the activated forms of monosaccharides, are used as donor substrates by glycosyltransferases in the in vivo biosynthesis of polysaccharides, proteoglycans and glycoglycerolipids. In mammals, uridine 5'-diphosphate-glucose (UDP-Glc) and uridine 5'-diphosphate-glucuronic acid (UDP-GlcA) are common building blocks of glycans and glycoconjugates. The commercial demand for these high-energy donors is increasing, while the understanding and application of enzymatic glycan synthesis has leapt forward in recent years. To produce valuable UDP-GlcA in a cost-effective way, here, UDP-sugar pyrophosphorylase from Arabidopsis thaliana was constitutively expressed in Pichia pastoris and secreted into the extracellular medium. Under these conditions, the synthesis of 4.2 g UDP-GlcA or 5.5 g UDP-Glc per liter of culture was revealed in the culture medium, without any need for purification. An anion exchange chromatography purification method for UDP-sugars was also developed. This route opens a door to large-scale production of the cheaper UDP-GlcA.