Abstract
N-Acetyl-D-neuraminic acid lyase (NanA) catalyzes the breakdown of sialic acid (Neu5Ac) to N-acetyl-D-mannosamine (ManNAc) and pyruvate. NanA plays a key role in Neu5Ac catabolism in many pathogenic and bacterial commensals where sialic acid is available as a carbon and nitrogen source. Several pathogens or commensals decorate their surfaces with sialic acids as a strategy to escape host innate immunity. Catabolism of sialic acid is key to a range of host-pathogen interactions. In this study, atomic resolution structures of NanA from Fusobacterium nucleatum (FnNanA) in ligand-free and ligand-bound forms are reported at 2.32 and 1.76 Å resolution, respectively. F. nucleatum is a Gram-negative pathogen that causes gingival and periodontal diseases in human hosts. Like other bacterial N-acetylneuraminate lyases, FnNanA also shares the triosephosphate isomerase (TIM)-barrel fold. As observed in other homologous enzymes, FnNanA forms a tetramer. In order to characterize the structure-function relationship, the steady-state kinetic parameters of the enzyme are also reported.
Highlights
Sialic acids are a family of related nine-carbon, acidic, -keto sugars which are used by bacteria for molecular mimicry
neuraminic acid lyase (NanA) belongs to the class I aldolase family, which is characterized by a triosephosphate isomerase (TIM)-barrel fold and by aldol condensation that proceeds through the formation of a Schiff base intermediate between a conserved lysine residue and the substrate pyruvate (Campeotto et al, 2009)
The sequence identities of the N-acetylneuraminate lyases from Haemophilus influenzae (PDB entry 1f7b; Barbosa et al, 2000), P. multocida (PDB entry 4imd; Huynh et al, 2013), Staphylococcus aureus (PDB entry 5a8g; Stockwell et al, 2016) and E. coli (PDB entry 1nal; Izard et al, 1994) are 74, 72, 57 and 36%, respectively
Summary
Sialic acids are a family of related nine-carbon, acidic, -keto sugars which are used by bacteria for molecular mimicry. There are five enzymes that are responsible for the catabolism of sialic acids (Fig. 1a). N-Acetylneuraminate lyase (NanA), the first committed enzyme, cleaves Neu5Ac to N-acetyl-d-mannosamine (ManNAc) and pyruvate (Fig. 1b). N-Acetyl-d-neuraminic acid lyase (NanA; EC 4.1.3.3) is known as sialic acid aldolase. NanA belongs to the class I aldolase family, which is characterized by a triosephosphate isomerase (TIM)-barrel fold and by aldol condensation that proceeds through the formation of a Schiff base intermediate between a conserved lysine residue and the substrate pyruvate (Campeotto et al, 2009). We report 2.32 and 1.76 Aresolution structures of N-acetylneuraminate lyase from F. nucleatum in two forms: ligand-free and with ligand bound as a pyruvate Schiff base. The high-resolution structure of the enzyme is compared with those of other known structures of sialic acid aldolases. We report the steady-state enzyme kinetics of this enzyme from F. nucleatum
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