tetano
Editor, Senior Moderator
N Biotechnol
. 2021 Feb 5;S1871-6784(21)00015-7.
doi: 10.1016/j.nbt.2021.01.010. Online ahead of print.
FN3-based Monobodies that Bind to the Receptor Binding Domain of the SARS-CoV-2 Spike Protein
Christina J Miller[SUP] 1 [/SUP], Jennifer E McGinnis[SUP] 1 [/SUP], Michael J Martinez[SUP] 1 [/SUP], Guangli Wang[SUP] 2 [/SUP], Jian Zhou[SUP] 3 [/SUP], Erica Simmons[SUP] 4 [/SUP], Tohti Amet[SUP] 4 [/SUP], Sanofar J Abdeen[SUP] 4 [/SUP], James W Van Huysse[SUP] 4 [/SUP], Ronald R Bowsher[SUP] 4 [/SUP], Brian K Kay[SUP] 5 [/SUP]
Affiliations
Abstract
A phage library displaying 10[SUP]10[/SUP] variants of the fibronectin type III (FN3) domain was affinity selected with the biotinylated form of the receptor binding domain (RBD, residues 319-541) of the SARS-CoV-2 virus spike protein. Nine binding FN3 variants (i.e. monobodies) were recovered, representing four different primary structures. Soluble forms of the monobodies bound to several different preparations of the RBD and the S1 spike subunit, with affinities ranging from 3 to 14 nM as measured by bio-layer interferometry. Three of the four monobodies bound selectively to the RBD of SARS-CoV-2, with the fourth monobody showing slight cross-reactivity to the RBD of SARS-CoV-1 virus. Examination of binding to the spike fragments and its trimeric form revealed that the monobodies recognise at least three overlapping epitopes on the RBD of SARS-CoV-2. While pairwise tests failed to identify a monobody pair that could bind simultaneously to the RBD, one monobody could simultaneously bind to the RBD and the ectodomain of the cellular receptor angiotensin converting enzyme 2 (ACE2). All four monobodies successfully bound the RBD after overexpression in Chinese hamster ovary (CHO) cells as fusions to the Fc domain of human IgG1.
Keywords: SARS-CoV-1 virus; SARS-CoV-2 virus; affinity selection; angiotensin-converting enzyme; coronavirus; monobody; phage-display; receptor; spike protein.
. 2021 Feb 5;S1871-6784(21)00015-7.
doi: 10.1016/j.nbt.2021.01.010. Online ahead of print.
FN3-based Monobodies that Bind to the Receptor Binding Domain of the SARS-CoV-2 Spike Protein
Christina J Miller[SUP] 1 [/SUP], Jennifer E McGinnis[SUP] 1 [/SUP], Michael J Martinez[SUP] 1 [/SUP], Guangli Wang[SUP] 2 [/SUP], Jian Zhou[SUP] 3 [/SUP], Erica Simmons[SUP] 4 [/SUP], Tohti Amet[SUP] 4 [/SUP], Sanofar J Abdeen[SUP] 4 [/SUP], James W Van Huysse[SUP] 4 [/SUP], Ronald R Bowsher[SUP] 4 [/SUP], Brian K Kay[SUP] 5 [/SUP]
Affiliations
- PMID: 33556628
- PMCID: PMC7863792
- DOI: 10.1016/j.nbt.2021.01.010
Abstract
A phage library displaying 10[SUP]10[/SUP] variants of the fibronectin type III (FN3) domain was affinity selected with the biotinylated form of the receptor binding domain (RBD, residues 319-541) of the SARS-CoV-2 virus spike protein. Nine binding FN3 variants (i.e. monobodies) were recovered, representing four different primary structures. Soluble forms of the monobodies bound to several different preparations of the RBD and the S1 spike subunit, with affinities ranging from 3 to 14 nM as measured by bio-layer interferometry. Three of the four monobodies bound selectively to the RBD of SARS-CoV-2, with the fourth monobody showing slight cross-reactivity to the RBD of SARS-CoV-1 virus. Examination of binding to the spike fragments and its trimeric form revealed that the monobodies recognise at least three overlapping epitopes on the RBD of SARS-CoV-2. While pairwise tests failed to identify a monobody pair that could bind simultaneously to the RBD, one monobody could simultaneously bind to the RBD and the ectodomain of the cellular receptor angiotensin converting enzyme 2 (ACE2). All four monobodies successfully bound the RBD after overexpression in Chinese hamster ovary (CHO) cells as fusions to the Fc domain of human IgG1.
Keywords: SARS-CoV-1 virus; SARS-CoV-2 virus; affinity selection; angiotensin-converting enzyme; coronavirus; monobody; phage-display; receptor; spike protein.