title: Mutation of conserved N-glycosylation sites around the CD4-binding site of human immunodeficiency virus type 1 GP120 affects viral infectivity.
abstract: Infection by the human immunodeficiency virus type 1 (HIV-1) is initiated through interaction of its exterior envelope glycoprotein gp120 with the CD4 receptor on target cells. To address the possible role of N-glycosylation of HIV-1 gp120 in binding CD4, we mutated different conserved N-glycosylation site Asn-residues in the vicinity of the putative CD4 binding site, as single mutations or in combinations. Authentic and mutant gp120 proteins were produced using the baculovirus expression system. All mutant proteins were produced and secreted at similar levels and could be immunoprecipitated with an HIV(+)-serum. Furthermore, all glycosylation mutants retained the full capacity to bind CD4 except for a triple mutant which showed reduced binding. Different gp120 mutant genes were then introduced in an infectious proviral DNA clone. Upon transfection of MT-2 cells, the authentic HIV-1 clone induced maximal virus production after 6 days. In the case of the triple glycosylation mutant, comparable virus production was first reached after a delay of about 12 days. Moreover, in contrast to native HIV, the mutant virus induced no typical multinucleated giant cells. These results suggest that the attached carbohydrates around the CD4-binding site of gp120, may contribute to the generation of this protein domain required for high affinity receptor interaction.
authors.affiliation: Central Research Units, F. Hoffmann-La Roche Ltd, Basle, Switzerland.
authors.forename: L
authors.initials: L
authors.lastname: Dirckx
authors.forename: D
authors.initials: D
authors.lastname: Lindemann
authors.forename: R
authors.initials: R
authors.lastname: Ette
authors.forename: C
authors.initials: C
authors.lastname: Manzoni
authors.forename: D
authors.initials: D
authors.lastname: Moritz
authors.forename: J
authors.initials: J
authors.lastname: Mous
mesh_terms: D000163:Acquired Immunodeficiency Syndrome / Q000235:genetics / Q000276:immunology*; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D015704:CD4 Antigens / Q000276:immunology*; D002478:Cells, Cultured; D015726:Giant Cells / Q000382:microbiology; D006031:Glycosylation; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000254:growth & development / Q000276:immunology*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D012995:Solubility; D014162:Transfection; D014779:Virus Replication
chemical_list: D015704:CD4 Antigens; D015699:HIV Envelope Protein gp120