title: Identification of residues in the receptor-binding domain (RBD) of the spike protein of human coronavirus NL63 that are critical for the RBD-ACE2 receptor interaction.
abstract: Human coronavirus NL63 (NL63), a member of the group I coronaviruses, may cause acute respiratory diseases in young children and immunocompromised adults. Like severe acute respiratory syndrome coronavirus (SARS-CoV), NL63 also employs the human angiotensin-converting enzyme 2 (hACE2) receptor for cellular entry. To identify residues in the spike protein of NL63 that are important for hACE2 binding, this study first generated a series of S1-truncated variants, examined their associations with the hACE2 receptor and subsequently mapped a minimal receptor-binding domain (RBD) that consisted of 141 residues (aa 476-616) towards the C terminus of the S1 domain. The data also demonstrated that the NL63 RBD bound to hACE2 more efficiently than its full-length counterpart and had a binding efficiency comparable to the S1 or RBD of SARS-CoV. A further series of RBD variants was generated using site-directed mutagenesis and random mutant library screening assays, and identified 15 residues (C497, Y498, V499, C500, K501, R518, R530, V531, G534, G537, D538, S540, E582, W585 and T591) that appeared to be critical for the RBD-hACE2 association. These critical residues clustered in three separate regions (designated RI, RII and RIII) inside the RBD, which may represent three receptor-binding sites. These results may help to delineate the molecular interactions between the S protein of NL63 and the hACE2 receptor, and may also enhance our understanding of the pathogenesis of NL63 and SARS-CoV.
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Han-Xin
authors.initials: HX
authors.lastname: Lin
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Yan
authors.initials: Y
authors.lastname: Feng
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Gillian
authors.initials: G
authors.lastname: Wong
authors.affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: Liping
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Bei
authors.initials: B
authors.lastname: Li
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Xuesen
authors.initials: X
authors.lastname: Zhao
authors.affiliation: National Microbiology Laboratory, Canadian Science Center for Human and Animal Health, Winnipeg, MB R3E 3R2, Canada.
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Fiona
authors.initials: F
authors.lastname: Smaill
authors.affiliation: Department of Pathology and Molecular Medicine, McMaster University and Department of Microbiology, St Joseph's Healthcare, Hamilton, ON L8N 4A6, Canada.
authors.forename: Chengsheng
authors.initials: C
authors.lastname: Zhang
mesh_terms: D000085962:Angiotensin-Converting Enzyme 2; D002460:Cell Line; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000502:physiology; D025941:Protein Interaction Mapping; D011991:Receptors, Virus / Q000378:metabolism*; D045473:Severe acute respiratory syndrome-related coronavirus / Q000737:chemistry* / Q000378:metabolism; D045169:Severe Acute Respiratory Syndrome / Q000821:virology; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D008562:Membrane Glycoproteins; D011991:Receptors, Virus; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus; D007703:Peptidyl-Dipeptidase A; C000705307:ACE2 protein, human; D000085962:Angiotensin-Converting Enzyme 2