Sialic acid transport in Haemophilus influenzae is essential for lipopolysaccharide sialylation and serum resistance and is dependent on a novel tripartite ATP-independent periplasmic transporter

TY - JOUR

T1 - Sialic acid transport in Haemophilus influenzae is essential for lipopolysaccharide sialylation and serum resistance and is dependent on a novel tripartite ATP-independent periplasmic transporter

AU - Severi, E

AU - Randle, G

AU - Kivlin, P

AU - Whitfield, K

AU - Young, R

AU - Moxon, R

AU - Kelly, D

AU - Hood, D

AU - Thomas, G H

PY - 2005/11

Y1 - 2005/11

N2 - Sialylation of the lipopolysaccharide (LPS) is an important mechanism used by the human pathogen Haemophilus influenzae to evade the innate immune response of the host. We have demonstrated that N-acetylneuraminic acid (Neu5Ac or sialic acid) uptake in H. influenzae is essential for the subsequent modification of the LPS and that this uptake is mediated through a single transport system which is a member of the tripartite ATP-independent periplasmic (TRAP) transporter family. Disruption of either the siaP (HI0146) or siaQM (HI0147) genes, that encode the two subunits of this transporter, results in a complete loss of uptake of [C-14]-Neu5Ac. Mutant strains lack sialylated glycoforms in their LPS and are more sensitive to killing by human serum than the parent strain. The SiaP protein has been purified and demonstrated to bind a stoichiometric amount of Neu5Ac by electrospray mass spectrometry. This binding was of high affinity with a K-d of similar to 0.1 mu M as determined by protein fluorescence. The inactivation of the SiaPQM TRAP transporter also results in decreased growth of H. influenzae in a chemically defined medium containing Neu5Ac, supporting an additional nutritional role of sialic acid in H. influenzae physiology.

AB - Sialylation of the lipopolysaccharide (LPS) is an important mechanism used by the human pathogen Haemophilus influenzae to evade the innate immune response of the host. We have demonstrated that N-acetylneuraminic acid (Neu5Ac or sialic acid) uptake in H. influenzae is essential for the subsequent modification of the LPS and that this uptake is mediated through a single transport system which is a member of the tripartite ATP-independent periplasmic (TRAP) transporter family. Disruption of either the siaP (HI0146) or siaQM (HI0147) genes, that encode the two subunits of this transporter, results in a complete loss of uptake of [C-14]-Neu5Ac. Mutant strains lack sialylated glycoforms in their LPS and are more sensitive to killing by human serum than the parent strain. The SiaP protein has been purified and demonstrated to bind a stoichiometric amount of Neu5Ac by electrospray mass spectrometry. This binding was of high affinity with a K-d of similar to 0.1 mu M as determined by protein fluorescence. The inactivation of the SiaPQM TRAP transporter also results in decreased growth of H. influenzae in a chemically defined medium containing Neu5Ac, supporting an additional nutritional role of sialic acid in H. influenzae physiology.

KW - ESCHERICHIA-COLI

KW - TRAP TRANSPORTERS

KW - STRUCTURAL-CHARACTERIZATION

KW - RHODOBACTER-CAPSULATUS

KW - GEL-ELECTROPHORESIS

KW - VIBRIO-CHOLERAE

KW - IDENTIFICATION

KW - GENES

KW - MEMBRANE

KW - METABOLISM

U2 - 10.1111/j.1365-2958.2005.04901.x

DO - 10.1111/j.1365-2958.2005.04901.x

M3 - Article

SN - 0950-382X

VL - 58

SP - 1173

EP - 1185

JO - Molecular Microbiology

JF - Molecular Microbiology

IS - 4

ER -