The capsular polysaccharides (CPS) of Campylobacter jejuni contain multiple heptose residues with variable stereochemical arrangements at C3−C6. The immediate precursor to all of these possible variations is currently believed to be GDP-D-glycero-α-Dmanno-heptose. Oxidation of this substrate at C4 enables subsequent epimerization reactions at C3−C5 that can be coupled to the dehydration/reduction at C5/C6. However, the enzyme responsible for the critical oxidation of C4 from GDP-D-glycero-α-D-mannoheptose has remained elusive. The enzyme Cj1427 from C. jejuni NCTC 11168 was shown to catalyze the oxidation of GDP-D-glyceroα-D-manno-heptose to GDP-D-glycero-4-keto-α-D-lyxo-heptose in the presence of α-ketoglutarate using mass spectrometry and nuclear magnetic resonance spectroscopy. At pH 7.4, the apparent k cat is 0.6 s −1 , with a value of k cat /K m of 1.0 × 10 4 M −1 s −1 for GDP-D-glycero-α-D-manno-heptose. α-Ketoglutarate is required to recycle the tightly bound NADH nucleotide in the active site of Cj1427, which does not dissociate from the enzyme during catalysis.