title: Introduction of N-linked glycans in the lectin domain of surfactant protein D: impact on interactions with influenza A viruses.
abstract: Porcine surfactant protein D (pSP-D) displays distinctively strong, broad-range inhibitory activity against influenza A virus (IAV). N-Linked glycosylation of the carbohydrate recognition domain (CRD) of pSP-D contributes to the high affinity of this collectin for IAV. To investigate the role of the N-linked glycan further, HEK293E protein expression was used to produce recombinant pSP-D (RpSP-D) that has similar structural and antiviral properties as NpSP-D. We introduced an additional N-linked glycan in the CRD of RpSP-D but this modification did not alter the antiviral activity. Human SP-D is unglycosylated in its CRD and less active against IAV compared with pSP-D. In an attempt to modify its antiviral properties, several recombinant human SP-D (RhSP-D) mutants were constructed with N-linked glycans introduced at various locations within its CRD. To retain lectin activity, necessary for the primary interactions between SP-D and IAV, N-linked glycosylation of RhSP-D was shown to be restricted to the corresponding position in the CRD of either pSP-D or surfactant protein A (SP-A). These N-glycosylated RhSP-D mutants, however, did not show increased neutralization activity against IAV. By developing RhSP-D mutants that also have the pSP-D-specific Ser-Gly-Ala loop inserted in the CRD, we could demonstrate that the N-linked glycan-mediated interactions between pSP-D and IAV involves additional structural prerequisites of the pSP-D CRD. Ultimately, these studies will help to develop highly effective SP-D-based therapeutic and prophylactic drugs against IAV.
authors.affiliation: Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Utrecht 3584CL, The Netherlands. m.vaneijk@uu.nl
authors.forename: Martin
authors.initials: M
authors.lastname: van Eijk
authors.forename: Laurie
authors.initials: L
authors.lastname: Bruinsma
authors.forename: Kevan L
authors.initials: KL
authors.lastname: Hartshorn
authors.forename: Mitchell R
authors.initials: MR
authors.lastname: White
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Rynkiewicz
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Seaton
authors.forename: Wieger
authors.initials: W
authors.lastname: Hemrika
authors.forename: Roland A
authors.initials: RA
authors.lastname: Romijn
authors.forename: Bas W
authors.initials: BW
authors.lastname: van Balkom
authors.forename: Henk P
authors.initials: HP
authors.lastname: Haagsman
mesh_terms: D000818:Animals; D004285:Dogs; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D009980:Influenza A virus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007251:Influenza, Human / Q000235:genetics / Q000628:therapy; D037102:Lectins; D009154:Mutation; D011134:Polysaccharides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D037662:Pulmonary Surfactant-Associated Protein A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D037663:Pulmonary Surfactant-Associated Protein D / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013552:Swine
chemical_list: D037102:Lectins; D011134:Polysaccharides; D037662:Pulmonary Surfactant-Associated Protein A; D037663:Pulmonary Surfactant-Associated Protein D; D011994:Recombinant Proteins