Identification and characterization of a deaminoneuraminic acid (Kdn)-specific aldolase from Sphingobacterium species.

Sialic acid (Sia) is a group of acidic sugars with a 9-carbon backbone, and classified into 3 species based on the substituent group at C5 position: N-acetylneuraminic acid (Neu5Ac), N-glycolylneuraminic acid (Neu5Gc), and deaminoneuraminic acid (Kdn). In Escherichia coli, the sialate aldolase or N-acetylneuraminate aldolase (NanA) is known to catabolize these Sia species into pyruvate and the corresponding 6-carbon mannose derivatives. However, in bacteria, very little is known about the catabolism of Kdn, compared with Neu5Ac. In this study, we found a novel Kdn-specific aldolase (Kdn-aldolase), which can exclusively degrade Kdn, but not Neu5Ac or Neu5Gc, from Sphingobacterium sp., which was previously isolated from a Kdn-assimilating bacterium. Kdn-aldolase had the optimal pH and temperature at 7.0-8.0 and 50 °C, respectively. It also had the synthetic activity of Kdn from pyruvate and mannose. Site-specific mutagenesis revealed that N50 residue was important for the Kdn-specific reaction. Existence of the Kdn-aldolase suggests that Kdn-specific metabolism may play a specialized role in some bacteria.

Identification and characterization of a novel, versatile sialidase from a Sphingobacterium that can hydrolyze the glycosides of any sialic acid species at neutral pH.

Bacterial sialidases are widely used to remove sialic acid (Sia) residues from glycans. Most of them cleave the glycosides of N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) under acidic pHs; however, currently available bacterial sialidases had no activity to the glycosides of deaminoneuraminic acid (Kdn). In this study, we found a novel sialidase from Sphingobacterium sp. strain HMA12 that could cleave any of the glycosides of Neu5Ac, Neu5Gc, and Kdn. It also had a broad linkage specificity, i.e., α2,3-, α2,6-, α2,8-, and α2,9-linkages, and the optimal pH at neutral ranges, pH 6.5-7.0. These properties are particularly important when sialidases are applied for in vivo digestion of the cell surface sialosides under physiological conditions. Interestingly, 2,3-didehydro-2-deoxy-N-acetylneuraminic acid (Neu5Ac2en), which is a transition state analog-based inhibitor, competitively inhibited the enzyme-catalyzed reaction for Kdn as well as for Neu5Ac, suggesting that the active site is common to the Neu5Ac and Kdn residues. Taken together, this sialidase is versatile and useful for the in vivo research on sialo-glycoconjugates.