Glycosylation Characterization of an Influenza H5N7 Hemagglutinin Series with Engineered Glycosylation Patterns: Implications for Structure-Function Relationships.
J Proteome Res
; 16(2): 398-412, 2017 02 03.
Article
en En
| MEDLINE
| ID: mdl-28060516
ABSTRACT
The glycosylation patterns of four recombinant H5 hemagglutinins (HAs) derived from A/Mallard/Denmark/64650/03 (H5N7) have been characterized. The proteins were expressed in (i) HEK293T cells to produce complex glycoforms, (ii) HEK293T cells treated with Vibrio cholera neuraminidase to provide asialo-complex glycoforms, (iii) HEK293S GnTI(-) cells with predominantly the canonical Man5GlcNAc2 glycoform, and (iv) Drosophila S2 insect cells producing primarily paucimannose glycoforms. Previously, these HAs were used to investigate the effect of different glycosylation states on the immune responses in chicken and mouse systems. Evidence was found that high-mannose glycans diminished antibody response via DC-SIGN interactions. We performed two semiquantitative analyses including MALDI-TOF MS permethylation analysis of released glycans and LC-MSE analysis of glycosylation site microheterogeneity. Glycosylation site occupancy was also determined by LC-MSE. Our major findings include (1) decreasing complexity of glycosylation from the stem to the globular head, (2) absence of glycosylation at N10 and N193, (3) complex glycans at N165 in HEK293T cell HA but high mannose glycans at this site in HEK293S and S2 cells, and (4) differences between the three-dimensional structures of H3 and H5 HAs that may explain glycan type preferences at selected sites. Biological implications of the findings are discussed.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Virus de la Influenza A
/
Ingeniería de Proteínas
/
Glicoproteínas Hemaglutininas del Virus de la Influenza
/
Manosa
Tipo de estudio:
Prognostic_studies
Límite:
Animals
/
Humans
Idioma:
En
Revista:
J Proteome Res
Asunto de la revista:
BIOQUIMICA
Año:
2017
Tipo del documento:
Article
País de afiliación:
Estados Unidos