The structure and interaction of spike protein and the receptor of SARS-Cov-2 (II)
Figure 2. The whole structure of the combination of SARS-CoV-2 RBD and ACE2
2.2 S2 subunit-mediated membrane fusion
The S1 subunit mediates receptor binding, and the S2 subunit is responsible for membrane fusion. Membrane fusion is the result of S protein activation, and depends on the cleavage of S protein at S1/S2 and S2' sites by host cell proteases (such as furin), which is an indispensable step for viruses to enter cells.
The S1/S2 site of SARS-CoV-2 is an exposed ring structure containing multiple arginine residues, which is very different from SARS-CoV. This structure is more conducive to furin cleavage. Furin cuts the arginine (Arg)-XY-arginine (Arg) in the peptide chain (X is any amino acid, Y is arginine or lysine). (Acid) peptide bond to the carboxy terminal of the sequence. The so-called S2' site of the S protein upstream of the FP is further cleaved by the host protease to promote irreversible conformational changes to activate the S protein for membrane fusion. Transmembrane protease serines2 (TMPRSS2) is an important mediator of S protein activation, helps viruses enter cells, and is a potential drug target.
The S2 subunits of SARS-CoV and SARS-CoV-2 have 89.8% sequence homology. After the RBD of the S protein S1 subunit binds to the ACE2 receptor on the target cell, the two heptapeptide repeats HR1 and HR2 of the S protein S2 subunit interact to form the fusion core of the 6-helix bundle (6-HB). The virus and the cell membrane are tightly integrated, facilitating fusion and infection.
The three HR1 domains of the S protein trimer form a parallel trimer coiled center, and the three HR2 domains around the center are twisted together in an anti-parallel manner. The interaction between these two domains is mainly a hydrophobic force. Each pair of adjacent HR1 helices forms a deep hydrophobic groove, which provides binding sites for the hydrophobic residues of the HR2 domain. Amino acid sequence alignment showed that the homology of SARS-CoV-2 and SARS-CoV in HR1 and HR2 regions was 92.6% and 100%, respectively. Compared with SARS-CoV, SARS-CoV-2 has eight amino acid residues in the fusion core region of the HR1 domain. Crystallographic analysis shows that this may help to enhance the interaction between HR1 and HR2. Stabilizing the 6-HB conformation of SARS-CoV-2 may lead to an increase in the infection ability of the new coronavirus.
The homology of the amino acid sequence of the SARS-CoV and SARS-CoV-2 fusion peptide is 93%. The S2 subunit starts from the fusion peptide (FP), and downstream is HR1 and HR2. FP is the membrane fusion functional element of the S protein. FP is a conserved short fragment (15–25 amino acid residues) in the S protein of the coronavirus family, which is mainly composed of hydrophobic amino acids such as glycine or alanine that is inserted into the host cell membrane to initiate fusion.
Membrane fusion is a multi-step process. After S protein is activated, FP will directly interact with the host cell membrane to promote fusion to form a fusion pore. The virus then injects genomic RNA into the host cell through the fusion pore to replicate and produce more viruses. Studies have found that Ca2+ helps FP achieve its function and promotes the infection of host cells by SARS-CoV-2.
3. Guidance for drug development
Vaccines are the ultimate means of preventing viral infections. The latest research shows that patients’ antibody levels against the new coronavirus have not declined in the four months after the infection is confirmed. This suggests that antibody vaccines can protect the body in human immunity and have a certain maintenance time. However, because the coronavirus is a single-stranded RAN, it is very easy to mutate during its transmission, which may reduce the protective effect of the Cov vaccine against some mutant viruses. Currently, there is no clear antiviral treatment strategy for COVID-19, and clinically, supportive treatment and symptomatic treatment are the mainstay. Drugs or molecular preparations that directly interfere with the interaction between the virus and the host receptor have broad practical prospects: such as blocking the cleavage site of the virus, blocking the specific binding of ACE2 and the virus S protein, and using cell protease inhibitors in combination. At the same time, it can increase the circulating ACE2 concentration in the body, and play the protective effect of ACE2 on organs.
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