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Coronavirus Overall Structure Elaborated
Coronavirus Overall Structure Elaborated
Above is the basic structure of COVID-19 from ProSci
Image Credit: https://www.prosci-inc.com/covid-19/
Image Credit: https://www.saccounty.net/COVID-19/Pages/default.aspx
Spike Protein (S): COVID-19 infects human cells by interacting with the angiotensin-converting enzyme 2 (ACE2 receptor). The SARS-CoV-2 Spike (S) glycoprotein binds to the cell ACE2 receptor on the cell membrane in order to enter the cell. The Spike Protein is the largest structure on the Coronavirus, giving it a "crown-like" and "spiky" appearance. The Spike Protein is very efficient in infecting the host cell because it is heavily glycosylated, contains an N-terminal signal sequence which instructs it to bind to the ACE2 receptor, and is cleaved into two sub-units, S1 and S2 (more on the Spike protein below).
Genetic Code (RNA): Coronaviruses are RNA viruses.
Nucleocapsid Protein (N): The nucleocapsid protein helps the virus bind to the replicase-transcriptase complex (RTC), finally leading to the packaging of the viral genome.
Envelope Protein (E): The envelope protein is not found in large quantities in the virus. Research shows that the envelope protein functions like a transmembrane protein, which facilitates ion transport, assembly of the virus, and release of the virus. This protein is essential for the development of the virus (pathogenesis).
Membrane Protein (M): The membrane protein is a major dimer (dimer: two identical molecules linked together) on the Coronavirus. This protein has two conformations, allowing membrane curvature.
Hemagglutinin-esterase dimer protein (HE): The HE protein is present in various betacoronaviruses. This protein enhances viral entry by binding sialic acids on surface Spike glycoproteins.
Sources (all of above information):
Habibzadeh P, Stoneman EK. The novel coronavirus: A bird's eye view. Int J Occup Environ Med 2020;11:65-71. doi: 10.15171/ijoem.2020.1921.
ProSci. "COVID-19 (SARS-CoV-2, 2019-nCoV) Antibodies," https://www.prosci-inc.com/covid-19/
Basic Spike Protein Structure of Coronaviruses
Basic Spike Protein Structure of Coronaviruses
The Spike Protein (S) is an important protein in all Coronaviruses because it is the structure which binds to cell membrane receptors.
The Spike Protein (S) is an important protein in all Coronaviruses because it is the structure which binds to cell membrane receptors.
The Spike Protein (S) is a large type I transmembrane glycoprotein that ranges from 1,160 amino acids to 1,400 amino acids, with 21 to 35 N-Glycosylation sites (highly glycosylated). Spike Proteins are trimers on the surface of Coronaviruses, which means that three proteins are in one complex. Their structuring creates a spiky "crown-like" appearance for the Coronavirus.
The S-Protein has two main features: (1) the receptor-binding domain or RBD, which is the structure that attaches to host cells, and the (2) cleavage site, which allows the virus to open and enter the host cell. Its ectodomain starts with the N-terminal domain (or S1 subunit) and ends with the C-terminal domain (or S2 subunit). The S1 subunit, in which the receptor-binding domain is incorporated, contains the instructions for receptor binding while the S2 subunit controls fusion with the cell membrane. The S1 subunit varies in sequence between different Coronaviruses and is responsible for recognizing and binding to host cell receptors. The variations in S1 subunit sequences contributes to diversity in the Coronaviridae family.
As mentioned above, another feature of the S-protein is its cleavage site. The S glycoprotein needs to be cleaved by cell proteases (protease: an enzyme which breaks up proteins into smaller polypeptides) to successfully facilitate viral fusion and entry into the cell. Typically, Coronaviruses, including some betacoronaviruses and all gammacoronaviruses, have a cleavage site between the S1 and S2 subunits. Cleavage typically occurs by a furin, a Golgi-resident host protease -own words. However, each Coronavirus has a different cleavage requirement; they require specific cell proteases for cleavage, have distinctive cleaving natures, and different cleaving locations. These factors increase diversity among Coronaviruses (explained on the right).
Sources (all of above information):
Scripps Research Institute. "COVID-19 coronavirus epidemic has a natural origin." ScienceDaily. ScienceDaily, 17 March 2020. <www.sciencedaily.com/releases/2020/03/200317175442.htm>.
Sino Biological. "Spike Protein/S Protein," https://www.sinobiological.com/research/virus/hcov-spike-protein-overview
Virology Blog. "Furin cleavage site in the SARS-CoV-2 coronavirus glycoprotein," http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/
Above are labels of the different features of the SARS-CoV Spike Protein and which residues are similar between SARS-CoV and SARS-CoV-2
Image Credit: From MDPI, https://www.mdpi.com/1999-4915/12/3/254/htm
Why is there Coronavirus Diversity?
Why is there Coronavirus Diversity?
The Spike Protein (S) is structurally and chemically different in every Coronavirus species. This variation further leads to Coronavirus evolution and diversity.
The Spike Protein (S) is structurally and chemically different in every Coronavirus species. This variation further leads to Coronavirus evolution and diversity.
The Spike Protein (S) has evolved and mutated into various structures, each with a unique receptor-binding interaction and cell membrane fusion pattern. Variations in the S-Protein structure, sequence, and cleavage play a big role in determining certain characteristics and distinguishing features in Coronaviruses including: (1) their receptor-binding interactions, and (2) their binding affinity, and (3) the severity of their infection on the body. Research shows that the receptor-binding domain (RBD) of different Coronaviruses is structurally different due to the distinctive S-Protein sequences and interactions with cell membrane surfaces and receptors (more on RBD structures below). Specifically, the S-protein of different Coronaviruses may either be cleaved or uncleaved during the assembly of virions, making it structurally different from all other viruses of its class. The cleavage requirements of Coronaviruses enhances diversity due to the differences in cell-virus interaction. For example: MHV-2 and MHV-A59 are two different strains of the betacoronavirus mouse hepatitis virus. These two strains have different cleavage requirements. As a result, their manner of fusing with the cell differs (fusogenecity). MERS-CoV has a furin cleavage site on its S-protein and is typically cleaved by intracellular proteases during exit from the cell. On the contrary, SARS-CoV is uncleaved when it is released from the cell but is cleaved during viral entry; this virus has two cleavage sites (as shown in the diagram below). Most alphacoronaviruses have an uncleaved S-protein when they are released from the cell. Intrestingly, proteolytic cleavage of the S glycoprotein plays a big role in determining whether a particular Coronavirus can pass from one species to another (e.g., bats to humans), also known as zoonotic Coronavirus transmission (read more about cleavage and zoonotic transmission below).
Sources (all of above information):
Sino Biological. "Spike Protein/S Protein," https://www.sinobiological.com/research/virus/hcov-spike-protein-overview
Wang, N., Shi, X., Jiang, L. et al. Structure of MERS-CoV spike receptor-binding domain complexed with human receptor DPP4. Cell Res 23, 986–993 (2013). https://doi.org/10.1038/cr.2013.92
Virology Blog. "Furin cleavage site in the SARS-CoV-2 coronavirus glycoprotein," http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/
Above is a representation of different betacornavirus S-Protein structures and the percentage of their amino acid similarities.
Image Credit: From bioRxiv, "Structural modeling of 2019-novel coronavirus (nCoV) spike protein reveals a proteolytically-sensitive activation loop as a distinguishing feature compared to SARS-CoV and related SARS-like coronaviruses," https://www.biorxiv.org/content/10.1101/2020.02.10.942185v1.full
Example of a Coronavirus Spike Protein Structure
Example of a Coronavirus Spike Protein Structure
Above is a representation of the S-protein of SARS-CoV. The ecotodomain of this Coronavirus is subdivided into the S1 subunit and the S2 subunit. The S1 subunit contains the receptor-binding domain and is responsible for binding to host cell receptors while the S2 subunit is responsible for fusion with the cell membrane. The S2 subunit contains the putative fusion peptide (blue) and the heptad repeat HR1 (orange) and HR2 (brown). The transmembrane domain is represented in purple - own words. Lastly, SARS-CoV has two cleavage sites, shown by the arrows in the diagram.
Image Credit: From Sino Biological, https://www.sinobiological.com/research/virus/hcov-spike-protein-overview
Spike Protein (S) in COVID-19
Spike Protein (S) in COVID-19
BACKGROUND: Binding to the ACE2 receptor
BACKGROUND: Binding to the ACE2 receptor
The Spike Protein (S) on COVID-19 is cleaved into two subunits, S1 and S2. While the S2 subunit enhances viral fusion with the cell membrane during viral entry, the S1 subunit contains the receptor-binding domain (RBD), allowing the Coronavirus to bind to the peptidase domain (PD) of the ACE2 receptor. The ACE2 receptor is typically bound to one of its ligands, amino acid transporter B0AT1 (amino acid transporter: transmembrane protein which transports amino acids). The RBD complex of COVID-19 therefore binds to the ACE2-B0AT1 complex in order to enter the cell. The Spike Protein on COVID-19 is a trimer (trimer: three Spike Proteins bound together). Research shows that two of the RBDs in the trimer face down while the third one faces up. The PD in the ACE2-B0AT1 complex can only bind to the RBD facing upward. However, the ACE2 receptor needs to dismerise (becoming a complex with two identical ACE2 molecules) in order for two Spike Protein trimers (three Spike Proteins bound together) to bind to the ACE2-B0AT1 dimer. Two Spike Proteins of the trimer will bind to the two PDs (peptidase domains) of the ACE2-B0AT1 dimer.
Source:
Drug Target Review. "Scientists demonstrate how COVID-19 infects human cells," https://www.drugtargetreview.com/news/56895/scientists-demonstrate-how-covid-19-infects-human-cells/
The structure above is the ACE2 receptor (in yellow) bound to the RBD in the COVID-19 Spike Protein (in magenta, turquoise, and green).
Image Credit: From Zhang Lab, https://zhanglab.ccmb.med.umich.edu/C-I-TASSER/2019-nCov/
Pre-fusion structure: COVID-19's Spike Protein trimer with three RBDs in total; two RBDs face down and one RBD faces up. The RBD downs are in white or gray and the RBD up is in green.
Image Credit: From Drug Target Review, https://www.drugtargetreview.com/news/56895/scientists-demonstrate-how-covid-19-infects-human-cells/
Image Credit: From Drug Target Review, https://www.drugtargetreview.com/news/56895/scientists-demonstrate-how-covid-19-infects-human-cells/
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SPIKE PROTEIN FEATURE #1: RBD (Receptor-Binding Domation)
SPIKE PROTEIN FEATURE #1: RBD (Receptor-Binding Domation)
COVID-19 v.s. SARS-CoV v.s. MERS-CoV
COVID-19:
The RBD of COVID-19 binds to the ACE2 receptor of humans, ferrets, cats and other homologous species with very high affinity. In fact, the binding affinity of COVID-19 is much higher than that of SARS-CoV yet it may bind less efficiently to the ACE2 receptors of organisms like rodents and civets, which are more prone to SARS-CoV-like viruses. L455, F486, Q493, S494, N501 and Y505 are six RBD amino acids which are critical for binding to ACE2 receptors. 5 of the 6 residues were not seen in SARS-CoV. Research shows that COVID-19 has a mutated RBD.
SARS-CoV:
The RBD of SARS-CoV also binds to the ACE2 receptor to enter the cell. However, unlike COVID-19, Y442, L472, N479, D480, T487 and Y4911 are the six RBD amino acids which are critical for binding to ACE2 receptors. Nevertheless, SARS-CoV still has a similar receptor-binding domain (RBD) structure to COVID-19 despite amino acid differences at 5 of the 6 key residues.
MERS-CoV:
The MERS-CoV Spike glycoprotein (S) RBD targets the dipeptidyl peptidase 4 (DPP4) receptor on the cell membrane, unlike SARS-CoV and SARS-CoV-2. Its RBD consists of a core and a receptor-binding subdomain. The RBD core of MERS-CoV is very similar to the RBD core of SARS-CoV. However, their receptor-binding subdomains have a lot of variation. This receptor-binding subdomain of MERS-CoV attaches to the DPP4 receptor. The DPP4 receptor is structurally different from the ACE2 receptor. Experiments show that the RBD binds to the extracellular domain of the DPP4 receptor, which consists of "N-terminal eight-bladed β-propeller domain (S39 to D496) and a C-terminal α/β hydrolase domain (N497 to P766). The β-propeller domain consists of eight blades, each made up of four antiparallel β-strands" - rephrase in own words. The RBD of MERS-CoV contacts blade four and five of the DPP4 receptor.
Sources (all of above information):
Andersen, K.G., Rambaut, A., Lipkin, W.I. et al. The proximal origin of SARS-CoV-2. Nat Med (2020). https://doi.org/10.1038/s41591-020-0820-9.
Habibzadeh P, Stoneman EK. The novel coronavirus: A bird's eye view. Int J Occup Environ Med 2020;11:65-71. doi: 10.15171/ijoem.2020.1921.
The Lancet. "Genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding," https://www.thelancet.com/journals/lancet/article/PIIS0140-6736(20)30251-8/fulltext#seccestitle10
Wang, N., Shi, X., Jiang, L. et al. Structure of MERS-CoV spike receptor-binding domain complexed with human receptor DPP4. Cell Res 23, 986–993 (2013). https://doi.org/10.1038/cr.2013.92
Above is a picture comparing the RBD (receptor-binding domain) sequences of COVID-19, SARS-CoV, and MERS-CoV. Click here for the detailed caption.
Image Credit: From Nature Articles: Cellular and Molecular Immunology, https://www.nature.com/articles/s41423-020-0400-4/figures/1
SPIKE PROTEIN FEATURE #2: Polybasic (furin) Cleavage Site & Predicted O-Linked Glycans
SPIKE PROTEIN FEATURE #2: Polybasic (furin) Cleavage Site & Predicted O-Linked Glycans
COVID-19 contains a polybasic cleavage site (RRAR) between the S1 and S2 subunits of the Spike Protein's ectodomain. Different from other Coronaviruses, the furin cleavage site of COVID-19 contains an inserted proline amino acid and its sequence is PRRARS|V. Turn in the proline amino acid has been predicted to to result in the addition of O-Linked Glycans in positions S673, T678 and S686 of the sequence. The effective cleavage of the COVID-19 Spike Protein by furin and other proteases enhances viral entry by strengthening the binding affinity. The Polybasic Cleavage Site and the predicted O-Linked Glycans have not been observed in other Lineage B betacoronaviruses but have been observed in HKU1, a Lineage A betacoronavirus. In fact, even the most homologous Coronavirus to COVID-19 isolated from a bat in Yunnan in 2013, RaTG-13, does not contain a furin cleavage site. COVID-19 most likely cleaves upon exit from epithelial cells since most of the furin proteases are present in the respiratory tract.
Protelytic cleavage of the S-protein can determine whether a particular Coronavirus will pass from one species to another (e.g., bats to human). Efficient cleavage of MERS-CoV gives it the potential to pass from bats to humans. A MERS-CoV from Ugandan bats can bind to the human cell. However, it cannot enter the cell due to the absence of the protease trypsin, which is necessary to enhance the cleavage of MERS-CoV so that it can enter the human cell. "This observation demonstrates that cleavage of the S glycoprotein is a barrier to zoonotic coronavirus transmission" (directly quoted from http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/). Another example is avian influenza A viruses, which can only enter the cell with cleavage of the HA glycoprotein. While low-pathogenic avian influenza viruses only contain a single basic amino acid in the cleavage site, high-pathogenic avian H5N1 influenza viruses contain a furin cleavage site in the HA glycoprotein. Virologists predict that the insertion of a furin cleavage site in COVID-19 allowed it to jump from bats to humans. They also say that the insertion of the furin cleavage site might have resulted from recombination with another virus that acquired this furin cleavage site. Their general conclusion about the furin cleavage site is as follows: "Acquisition of the furin cleavage site might be viewed as a ‘gain of function’ that enabled a bat CoV to jump into humans and begin its current epidemic spread" (directly quoted from http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/).
Sources (all of above information):
Andersen, K.G., Rambaut, A., Lipkin, W.I. et al. The proximal origin of SARS-CoV-2. Nat Med (2020). https://doi.org/10.1038/s41591-020-0820-9.
Virology Blog. "Furin cleavage site in the SARS-CoV-2 coronavirus glycoprotein," http://www.virology.ws/2020/02/13/furin-cleavage-site-in-the-sars-cov-2-coronavirus-glycoprotein/
Feature #1 (RBD) and #2 (Cleavage Site and O-Linked Glycans) summarized below.
Feature #1 (RBD) and #2 (Cleavage Site and O-Linked Glycans) summarized below.
Image Credit: From Nature Medicine, https://doi.org/10.1038/s41591-020-0820-9
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