title: Molecular and biological characterization of human monoclonal antibodies binding to the spike and nucleocapsid proteins of severe acute respiratory syndrome coronavirus.
abstract: Human monoclonal antibodies (MAbs) were selected from semisynthetic antibody phage display libraries by using whole irradiated severe acute respiratory syndrome (SARS) coronavirus (CoV) virions as target. We identified eight human MAbs binding to virus and infected cells, six of which could be mapped to two SARS-CoV structural proteins: the nucleocapsid (N) and spike (S) proteins. Two MAbs reacted with N protein. One of the N protein MAbs recognized a linear epitope conserved between all published human and animal SARS-CoV isolates, and the other bound to a nonlinear N epitope. These two N MAbs did not compete for binding to SARS-CoV. Four MAbs reacted with the S glycoprotein, and three of these MAbs neutralized SARS-CoV in vitro. All three neutralizing anti-S MAbs bound a recombinant S1 fragment comprising residues 318 to 510, a region previously identified as the SARS-CoV S receptor binding domain; the nonneutralizing MAb did not. Two strongly neutralizing anti-S1 MAbs blocked the binding of a recombinant S fragment (residues 1 to 565) to SARS-CoV-susceptible Vero cells completely, whereas a poorly neutralizing S1 MAb blocked binding only partially. The MAb ability to block S1-receptor binding and the level of neutralization of the two strongly neutralizing S1 MAbs correlated with the binding affinity to the S1 domain. Finally, epitope mapping, using recombinant S fragments (residues 318 to 510) containing naturally occurring mutations, revealed the importance of residue N479 for the binding of the most potent neutralizing MAb, CR3014. The complete set of SARS-CoV MAbs described here may be useful for diagnosis, chemoprophylaxis, and therapy of SARS-CoV infection and disease.
authors.affiliation: Crucell Holland BV, PO Box 2048, 2301 CA Leiden, The Netherlands.
authors.forename: Edward N
authors.initials: EN
authors.lastname: van den Brink
authors.forename: Jan
authors.initials: J
authors.lastname: Ter Meulen
authors.forename: Freek
authors.initials: F
authors.lastname: Cox
authors.forename: Mandy A C
authors.initials: MA
authors.lastname: Jongeneelen
authors.forename: Alexandra
authors.initials: A
authors.lastname: Thijsse
authors.forename: Mark
authors.initials: M
authors.lastname: Throsby
authors.forename: Wilfred E
authors.initials: WE
authors.lastname: Marissen
authors.forename: Pauline M L
authors.initials: PM
authors.lastname: Rood
authors.forename: Alexander B H
authors.initials: AB
authors.lastname: Bakker
authors.forename: Hans R
authors.initials: HR
authors.lastname: Gelderblom
authors.forename: Byron E
authors.initials: BE
authors.lastname: Martina
authors.forename: Albert D M E
authors.initials: AD
authors.lastname: Osterhaus
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Preiser
authors.forename: Hans Wilhelm
authors.initials: HW
authors.lastname: Doerr
authors.forename: John
authors.initials: J
authors.lastname: de Kruif
authors.forename: Jaap
authors.initials: J
authors.lastname: Goudsmit
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D000914:Antibodies, Viral / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000918:Antibody Specificity; D001665:Binding Sites; D002522:Chlorocebus aethiops; D000086462:Coronavirus Nucleocapsid Proteins; D018604:Epitope Mapping; D006801:Humans; D008562:Membrane Glycoproteins / Q000276:immunology*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D019590:Nucleocapsid Proteins / Q000276:immunology*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000276:immunology*; D064370:Spike Glycoprotein, Coronavirus; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000086462:Coronavirus Nucleocapsid Proteins; D008562:Membrane Glycoproteins; D019590:Nucleocapsid Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV