Journal: PLoS Pathogens

Article Title: The 2 nd sialic acid-binding site of influenza A virus neuraminidase is an important determinant of the hemagglutinin-neuraminidase-receptor balance

doi: 10.1371/journal.ppat.1007860

Figure Lengend Snippet: (A) Specific activity of hN2 and aN2 was determined by MUNANA assay and ELLA using different glycoprotein-lectin combinations (Fetuin-ECA, Fetuin-PNA, Fetuin-MAL I, Fetuin-SNA, Transferrin-ECA and Transferrin-SNA) and normalized to the specific activity of hN2 for MUNANA and each glycoprotein-lectin combination. (B) Specific activity of hN2 and aN2 is graphed normalized to the specific activity as determined for each protein by the Fetuin-ECA combination. Mean values and standard deviations from two independent experiments performed in triplicate are shown. Stars depict P values calculated using an unpaired two-tailed Student t test (**, P<0.01; ***, P<0.001). (C and D) BLI kinetic assay of NA enzymatic activity. Streptavidin biosensors were coated with biotinylated synthetic glycans (3’SLNLN, 6’SLNLN or LNLN). Subsequently, the sensors were incubated in buffer containing 4 μg aN2 or hN2 in the absence or presence of 8 μg ECA or ECA alone. ECA binding to sensors coated with 3’SLNLN or 6’SLNLN is a measure for SIA cleavage from these receptors by NA. Experiments were independently performed three times. Representative experiments are shown.

Article Snippet: After overnight incubation at 37°C, plates were washed and incubated with either biotinylated Erythrina Cristagalli Lectin (ECA, 1.25 μg/ml; Vector Laboratories), biotinylated peanut agglutinin (PNA, 2.5 μg/ml; Galab Technologied), biolinylated Sambucus Nigra Lectin (SNA, 1.25 μg/ml; Vector Laboratories) or biotinylated Maackia Amurensis Lectin I (MAL I, 2.5 μg/ml; Vector Laboratories).

Techniques: Activity Assay, Two Tailed Test, Kinetic Assay, Incubation, Binding Assay