The structure of the cyclic enterobacterial common antigen (ECA) from Yersinia pestis

Two antigenic acidic polysaccharides related to enterobacterial common antigen (ECA) were isolated from a vaccine strain of a pathogenic microorganism Yersinia pestis. The low molecular weight polysaccharide (LMP) is composed of equal amounts of 2-acetamido-2-deoxy- d-mannuronic acid, 4-acetamido-4,6-dideoxy- d-galactose (Fuc4NAc), and 2-amino-2-deoxy- d-glucose which is partially N- and partially 6- O-acetylated. The structure of the trisaccharide repeating unit was established by analyses of LMP and the completely N-acetylated LMP (LMP-NAc) using 1H and 13C NMR spectroscopy, including 2D COSY and 1D NOE spectroscopy. Deamination of LMP with nitrous acid gave a set of oligomers terminated with 2,5-anhydromannose and ranging from tri- to dodeca-saccharides, thus indicating a random distribution of free amino groups. FABMS analyses of LMP and LMP-NAc showed that LMP consists mainly of the cyclic tetramer of the trisaccharide repeating unit together with a small amount of the cyclic trimer and a very small amount of the cyclic pentamer and has, thus, the following structure: ▪ where R is Ac or H (≈ 1:1), R′ is Ac or H (≈ 1:4), and n = 4 (major), 3, 5 (minor). Small proportions of the linear trimer and the linear tetramer were also detected in the preparations. The high molecular weight polysaccharide is linear and has the same (or a very similar) repeating unit as LMP.