Product Specificity of C4-Reductases in the Biosynthesis of GDP-6-Deoxy-Heptoses during Capsular Polysaccharide Formation in Campylobacter jejuni .

Campylobacter jejuni is the leading cause of food poisoning in the United States and Europe. A capsular polysaccharide that coats the exterior of the bacterium helps evade the host immune system. At least 33 different strains of C. jejuni have been identified, and the chemical structures of 12 different capsular polysaccharides (CPSs) have been characterized from various serotypes. Thus far, 10 different heptose sugars have been found in the chemically characterized CPSs, and each of these are currently thought to originate from the modification of GDP-d- glycero -d- manno -heptose by the successive action of 4,6-dehydratase (or C4-dehydrogenase), C3- or C3/C5-epimerase, and C4-reductase. Within the sequenced strains of C. jejuni , we have identified 25 different C4-reductases that cluster into nine groups at a sequence identity of >90%. Eight of the proteins from seven different clusters were purified, and their product profiles were determined with GDP-6-deoxy-4-keto-heptose substrates using NMR and ESI mass spectrometry. The isolated products included GDP-6-deoxy-l- gluco -heptose (serotype HS:2), GDP-6-deoxy-l- galacto -heptose (serotype HS:42), GDP-6-deoxy-l- gulo -heptose (serotype HS:15), GDP-6-deoxy-d- ido -heptose (serotypes HS:3, HS:4, and HS:33), GDP-6-deoxy-d- manno -heptose (serotype HS:53), and GDP-6-deoxy-d- altro -heptose (serotype HS:23/36). Based on these observations, the product specificity can be reliably predicted for 14 additional C4-reductases from C. jejuni . The remaining three C4-reductases are highly likely to be required for the biosynthesis of 3,6-dideoxy-heptose products.