Taylor SARS-CoV2 Protease

SARS-CoV-2 is a positive-stranded RNA virus with nucleocapsid which belongs to betacoronaviruses. The 30 kb long +ssRNA contains a 5’-cap structure and a 3’-poly-A tail. After membrane fusion, the viral +ssRNA is released into the cytoplasm and translated into two polyproteins pp1a and pp1ab. ^{[1] [2]} Papain-like protease(s) and the main protease (also called 3C-like protease 3CLpro) are essential for processing the two polyproteins pp1a and pp1ab.

The coronavirus ORF 1 polyprotein can be divided into an N-terminal region that is processed by one or two Papain-like proteases and a C-terminal region which is processed by the main protease. ^[3] While papain-like protease(s) cleave only three sites, the main protease cleaves 11 sites in the polyprotein to generate functional proteins. Additionally, the main protease cleaves its own N- and C-terminal autoprocessing sites. The cleaved functional proteins include viral enzymes needed for replication such as the RNA-dependant RNA polymerase, a helicase and other non-structural or accessory proteins such as an exoribonuclease, an endoribonuclease, a ssRNA binding protein and a 2’-O-ribose methyltransferase. ^[4]

Overall Structure and Active Centre of 3CLpro

The main protease is a cysteine protease that is essential for the viral life cycle. It is folded like an augmented serine-protease which forms a homodimer consisting of the perpendicular protomers A and B. One protomer consists of three domains. Domain I and II (N-terminal domain) form an antiparallel chymotrypsin-like ß-barrel structure. Domain III (C-terminal end) consist of five alpha-helices arranged in an antiparallel cluster. ^{[5] [6]} For maximal protease activity, the protease forms a homodimer as the substrate binding site is located in a catalytic cleft between the two N-terminal ß-barrel structures (between domain I and II). The substrate binding site involves a catalytic dyad consisting of the residues Cys145 and His41. The N- and C-terminal domains are connected by a long loop. ^[7] N-terminal residues of each protomer which are called N-finger, make contact between the N- and C-terminal domains of the other protomer and thus are necessary for dimerization. ^[8] S1 is a substrate binding subsite pocket which lies next to the catalytic dyad and consists of the side chains Phe 140, His 163 and the main chains of Glu166, Asn142, Gly 143 and His172. It confers absolute specificity for the Gln-P1 substrate residue on the enzyme as the carbonyl oxygen of Gln-P1 is stabilized by an oxyanion hole which is formed by amide groups of Gly143 and the catalytic Cys145. ^{[9] [10]} Hence, polyproteins are cleaved within the Leu-Gln↓(Ser, Ala, Gly) sequence. ^[11]

3CLpro as Potential Drug Target

Due to a new outbreak of pulmonary diseases caused by SARS-CoVid-2, the development of new drugs is essential for containment of the viral spread. One promising drug target among coronaviruses is the main protease, as it is essential for processing the polyproteins translated from the viral RNA. Inhibiting this enzyme would block the viral replication and is unlikely to be toxic, as no human proteases with similar cleavage specificity are known. ^[12] The potential inhibitor classes can be divided into two classes based on their chemical structures. The first class involves peptide chains that fit the catalytic site of the enzyme by making a covalent link with Cys145, therefore blocking substrate binding. The second class consists of small organic compounds that bind the active site and hence act as competitive inhibitors. Thus, the substrate can not enter the active site cavity. A potential drug which belongs to the second class is Lopinavir, a HIV1 protease inhibitor which seems to be a promising candidate for the treatment of coronavirus infections. ^[13] </SX>

References

1. ↑ Guo, Y.-R., Cao, Q.-D., Hong, Z.-S., Tan, Y.-Y., Chen, S.-D., Jin, H.-J., Tan, K.-S., Wang, D.-Y. & Yan, Y. (2020). Mil Med Res. 7.
2. ↑ Cascella, M., Rajnik, M., Cuomo, A., Dulebohn, S. C. & Di Napoli, R. (2020). StatPearls, Vol. p. Treasure Island (FL): StatPearls Publishing.
3. ↑ Enjuanes, L., (2005). Coronavirus replication and reverse genetics Berlin; New York: Springer, S. 69-78.
4. ↑ Muramatsu, T., Takemoto, C., Kim, Y.-T., Wang, H., Nishii, W., Terada, T., Shirouzu, M. & Yokoyama, S. (2016). Proc Natl Acad Sci U S A. 113, 12997–13002.
5. ↑ Yang, H., Yang, M., Ding, Y., Liu, Y., Lou, Z., Zhou, Z., Sun, L., Mo, L., Ye, S., Pang, H., Gao, G. F., Anand, K., Bartlam, M., Hilgenfeld, R. & Rao, Z. (2003). Proc Natl Acad Sci U S A. 100, 13190–13195.
6. ↑ Xu, T., Ooi, A., Lee, H. C., Wilmouth, R., Liu, D. X. & Lescar, J. (2005). Acta Crystallogr Sect F Struct Biol Cryst Commun. 61, 964–966.
7. ↑ Anand, K., Ziebuhr, J., Wadhwani, P., Mesters, J. R. & Hilgenfeld, R. (2003). Science. 300, 1763–1767.
8. ↑ Yang, H., Xie, W., Xue, X., Yang, K., Ma, J., Liang, W., Zhao, Q., Zhou, Z., Pei, D., Ziebuhr, J., Hilgenfeld, R., Yuen, K. Y., Wong, L., Gao, G., Chen, S., Chen, Z., Ma, D., Bartlam, M. & Rao, Z. (2005). PLoS Biol. 3.
9. ↑ Gorbalenya, A. E., Snijder, E. J. & Ziebuhr, J. (2000). Journal of General Virology. 81, 853–879.
10. ↑ Xue, X., Yu, H., Yang, H., Xue, F., Wu, Z., Shen, W., Li, J., Zhou, Z., Ding, Y., Zhao, Q., Zhang, X. C., Liao, M., Bartlam, M. & Rao, Z. (2008). Journal of Virology. 82, 2515–2527.
11. ↑ Rut, W., Groborz, K., Zhang, L., Sun, X., Zmudzinski, M., Hilgenfeld, R. & Drag, M. (2020). BioRxiv. 2020.03.07.981928.
12. ↑ Zhang, L., Lin, D., Sun, X., Curth, U., Drosten, C., Sauerhering, L., Becker, S., Rox, K. & Hilgenfeld, R. (2020). Science.
13. ↑ Dayer, M. R., Taleb-Gassabi, S. & Dayer, M. S. (2017). Lopinavir; A Potent Drug against Coronavirus Infection: Insight from Molecular Docking Study.