Unexpected Diversity of Escherichia coli Sialate O-Acetyl Esterase NanS

Ariel Rangel, Susan M. Steenbergen, Eric R. Vimr

The sialic acids (N-acylneuraminates) are a group of nine-carbon keto-sugars existing mainly as terminal residues on animal glycoprotein and glycolipid carbohydrate chains. Bacterial commensals and pathogens exploit host sialic acids for nutrition, adhesion, or anti-recognition, where N-acetyl- or N-glycolylneuraminic acids are the two predominant chemical forms of sialic acids. Each form may be modified by acetyl esters at carbon positions 4, 7, 8, or 9, and by a variety of less common modifications. Modified sialic acids produce challenges for colonizing bacteria because the chemical alterations to Neu5Ac confer increased resistance to sialidase and aldolase activities essential for catabolism of host sialic acids. Bacteria with O-acetyl sialate esterase(s) utilize acetylated sialic acids for growth thereby gaining a presumed metabolic advantage over competitors lacking this activity. Here we demonstrate the esterase activity of E. coli NanS after purifying it as a C-terminal HaloTag™ fusion. Using a similar approach, we show that E. coli strain O157:H7 Stx prophage or prophage remnants invariably include paralogs of nanS often located downstream of the Shiga-like toxin genes. These paralogs may include sequences encoding N- or C-terminal domains of unknown function, but where the NanS domains can function as sialate O-acetyl esterases, as shown by complementation of an E. coli strain K-12 nanS mutant and the unimpaired growth of an E. coli O157 nanS mutant on O-acetylated sialic acid. We further demonstrate that nanS homologs in Streptococcus spp also encode active esterase, demonstrating an unexpected diversity of bacterial sialate O-acetyl esterase. The sialic acids are a family of over 40 naturally occurring 9-carbon keto-sugars that function in a variety of host-bacterial interactions. These sugars occur primarily as terminal carbohydrate residues on host glycoproteins and glycolipids. Available evidence indicates diverse bacterial species use host sialic acids for adhesion or as sources of carbon and nitrogen. Our results show that catabolism of the diacetylated form of host sialic acid requires a specialized esterase, NanS. Our results further show that nanS homologs exist in bacteria other than Escherichia coli, as well as part of toxigenic E. coli prophage. The unexpected diversity of these enzymes suggests new avenues for investigating host-bacterial interactions. Therefore, these original results extend our previous studies of nanS to include mucosal pathogens, prophage, and prophage remnants. This expansion of the nanS superfamily suggests important, though as yet unknown, functions in host-microbe interactions.