tetano
Editor, Senior Moderator
J Biol Chem. 2018 May 16. pii: jbc.RA117.001430. doi: 10.1074/jbc.RA117.001430. [Epub ahead of print]
[h=1]Lectin-mediated binding and sialoglycans of porcine surfactant protein D synergistically neutralize influenza A virus.[/h] van Eijk M[SUP]1[/SUP], Rynkiewicz MJ[SUP]2[/SUP], Khatri K[SUP]3[/SUP], Leymarie N[SUP]4[/SUP], Zaia J[SUP]2[/SUP], White MR[SUP]4[/SUP], Hartshorn KL[SUP]4[/SUP], Cafarella TR[SUP]2[/SUP], Van Die I[SUP]5[/SUP], Hessing M[SUP]6[/SUP], Seaton BA[SUP]7[/SUP], Haagsman HP[SUP]8[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Innate immunity is critical in the early containment of influenza A virus (IAV) infection, and surfactant protein D (SP-D) plays a crucial role in the pulmonary defense against IAV. In pigs, which are important intermediate hosts during the generation of pandemic IAVs, SP-D uses its unique carbohydrate recognition domain (CRD) to interact with IAV. An N-linked CRD-glycosylation provides interactions with the sialic acid binding site of IAV, and a tripeptide loop at the lectin binding site facilitates enhanced interactions with IAV glycans. Here, to investigate both mechanisms of IAV neutralization in greater detail, we produced an N-glycosylated neckCRD fragment of porcine SP-D (RpNCRD) in HEK293 cells. X-ray crystallography disclosed that the N-glycan did not alter the CRD backbone structure including the lectin site conformation, but revealed a potential second non-lectin binding site for glycans. IAV hemagglutination inhibition, IAV aggregation and neutralization of IAV infection studies showed that RpNCRD, unlike the human analogue RhNCRD, exhibits potent neutralizing activity against pandemic A/Aichi/68 (H3N2), enabled by both porcine-specific structural features of its CRD. MS analysis revealed an N-glycan site-occupancy of >98% at Asn303 of RpNCRD with complex-type, heterogeneously branched and predominantly α(2,3) sialylated oligosaccharides. Glycan binding array data characterized both RpNCRD and RhNCRD as mannose-type lectins. RpNCRD also bound LewisY structures whereas RhNCRD bound polylactosamine-containing glycans. Presence of the N-glycan in the CRD increases the glycan binding specificity of RpNCRD. These insights increase our understanding of porcine-specific innate defense against pandemic IAV and may inform the design of recombinant SP-D-based antiviral drugs.
[h=4]KEYWORDS:[/h] N-linked glycosylation; antiviral agent; collectin; drug design; host-pathogen interaction; influenza virus; innate immunity; porcine immunology; pulmonary defense; recombinant protein; sialic acid; structural biology; surfactant protein D; viral immunology
PMID: 29769321 DOI: 10.1074/jbc.RA117.001430
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[h=1]Lectin-mediated binding and sialoglycans of porcine surfactant protein D synergistically neutralize influenza A virus.[/h] van Eijk M[SUP]1[/SUP], Rynkiewicz MJ[SUP]2[/SUP], Khatri K[SUP]3[/SUP], Leymarie N[SUP]4[/SUP], Zaia J[SUP]2[/SUP], White MR[SUP]4[/SUP], Hartshorn KL[SUP]4[/SUP], Cafarella TR[SUP]2[/SUP], Van Die I[SUP]5[/SUP], Hessing M[SUP]6[/SUP], Seaton BA[SUP]7[/SUP], Haagsman HP[SUP]8[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] Innate immunity is critical in the early containment of influenza A virus (IAV) infection, and surfactant protein D (SP-D) plays a crucial role in the pulmonary defense against IAV. In pigs, which are important intermediate hosts during the generation of pandemic IAVs, SP-D uses its unique carbohydrate recognition domain (CRD) to interact with IAV. An N-linked CRD-glycosylation provides interactions with the sialic acid binding site of IAV, and a tripeptide loop at the lectin binding site facilitates enhanced interactions with IAV glycans. Here, to investigate both mechanisms of IAV neutralization in greater detail, we produced an N-glycosylated neckCRD fragment of porcine SP-D (RpNCRD) in HEK293 cells. X-ray crystallography disclosed that the N-glycan did not alter the CRD backbone structure including the lectin site conformation, but revealed a potential second non-lectin binding site for glycans. IAV hemagglutination inhibition, IAV aggregation and neutralization of IAV infection studies showed that RpNCRD, unlike the human analogue RhNCRD, exhibits potent neutralizing activity against pandemic A/Aichi/68 (H3N2), enabled by both porcine-specific structural features of its CRD. MS analysis revealed an N-glycan site-occupancy of >98% at Asn303 of RpNCRD with complex-type, heterogeneously branched and predominantly α(2,3) sialylated oligosaccharides. Glycan binding array data characterized both RpNCRD and RhNCRD as mannose-type lectins. RpNCRD also bound LewisY structures whereas RhNCRD bound polylactosamine-containing glycans. Presence of the N-glycan in the CRD increases the glycan binding specificity of RpNCRD. These insights increase our understanding of porcine-specific innate defense against pandemic IAV and may inform the design of recombinant SP-D-based antiviral drugs.
[h=4]KEYWORDS:[/h] N-linked glycosylation; antiviral agent; collectin; drug design; host-pathogen interaction; influenza virus; innate immunity; porcine immunology; pulmonary defense; recombinant protein; sialic acid; structural biology; surfactant protein D; viral immunology
PMID: 29769321 DOI: 10.1074/jbc.RA117.001430
Free full text