dbCAN-PUL

PUL ID	PUL0288
PubMed	22975275, Carbohydr Res. 2012 Oct 1;360:19-24. doi: 10.1016/j.carres.2012.07.013. Epub 2012 Jul 27.
Characterization method	acid hydrolysis,NMR,mass spectrometry,sugar utilization assay
Genomic accession number	X77617.1
Nucelotide position range	226-7140
Substrate	capsule polysaccharide,heparosan
Loci	kfiABCD
Species	Escherichia coli/562
Degradation or Biosynthesis	biosynthesis

PUL ID	PUL0288
PubMed	22975275, Carbohydr Res. 2012 Oct 1;360:19-24. doi: 10.1016/j.carres.2012.07.013. Epub 2012 Jul 27.
Title	Production of intracellular heparosan and derived oligosaccharides by lyase expression in metabolically engineered E. coli K-12.
Author	Barreteau H, Richard E, Drouillard S, Samain E, Priem B
Abstract	The cluster of genes of capsular K5 heparosan is composed of three regions, involved in the synthesis and the exportation of the polysaccharide. The region 2 possesses all the necessary genes involved in the synthesis of heparosan, namely kfiA, encoding alpha-4-N-acetylglucosaminyltransferase, kfiD, encoding beta-3-glucuronyl transferase, kfiC, encoding UDP-glucose dehydrogenase (UDP-glucuronic acid synthesis), and kfiB encoding a protein of unknown function. The cloning and expression of kfiADCB into Escherichia coli K-12 were found to be sufficient for the production of heparosan, which accumulates in the cells due to a lack of the exporting system. The concentration of recombinant heparosan reached one gram per liter under fed-batch cultivation. The cytoplasmic localization of heparosan inside the bacteria allowed subsequent enzymatic modifications such as a partial degradation with K5 lyase when expressed intracellularly. Under these conditions, the production of DP 2-10 oligosaccharides occurred intracellularly, at a concentration similar to that of recombinant intracellular heparosan.