Difference between revisions of "Part:BBa K5131000"

Revision as of 16:55, 1 October 2024

SARA-Cov-2 nsp5

Non-structural protein 5 (nsp5) is the most highly conserved non-structural protein across the Coronaviridae family, with a molecular weight of approximately 33 kDa[1].In SARS-Cov-2, it's referred to as the main protease due to its pivotal role in the cleavage of polyproteins. Nsp5 has up to 11 conserved cleavage sites on the polyprotein and first undergoes autolytic cleavage to release itself, after which it cleaves at these sites to release other non-structural proteins. Nsp5 consists of three structural domains: two N-terminal domains with protease activity and a C-terminal domain composed of α-helices. The active form of SARS-CoV-2 nsp5 is a homodimer that recognizes substrates approximately 10 amino acid residues in length, but exhibits selectivity at only four specific positions[2]. In addition to cleaving viral proteins, nsp5 also suppresses the host innate immune response by degrading host protein factors[3]. In this project, our primary goal is to express functional nsp5 in E.coli BL21 in preparation for in vivo inhibitor screening. Additionally, since nsp5 has the ability to recognize and cleave specific sequences, we also aim to develop it as a tool enzyme for removing recombinant tags during protein purification. Reference:

1. Jin Z, Du X, Xu Y, Deng Y, Liu M, Zhao Y, Zhang B, Li X, Zhang L, Peng C, Duan Y, Yu J, Wang L, Yang K, Liu F, Jiang R, Yang X, You T, Liu X, Yang X, Bai F, Liu H, Liu X, Guddat LW, Xu W, Xiao G, Qin C, Shi Z, Jiang H, Rao Z, Yang H. Structure of M(pro) from SARS-CoV-2 and discovery of its inhibitors. Nature. 2020 Jun;582(7811):289-293. doi: 10.1038/s41586-020-2223-y. Epub 2020 Apr 9. PMID: 32272481. 2. Zhao Y, Zhu Y, Liu X, Jin Z, Duan Y, Zhang Q, Wu C, Feng L, Du X, Zhao J, Shao M, Zhang B, Yang X, Wu L, Ji X, Guddat LW, Yang K, Rao Z, Yang H. Structural basis for replicase polyprotein cleavage and substrate specificity of main protease from SARS-CoV-2. Proc Natl Acad Sci U S A. 2022 Apr 19;119(16):e2117142119. doi: 10.1073/pnas.2117142119. Epub 2022 Apr 5. PMID: 35380892; PMCID: PMC9172370. 3.Wu Y, Ma L, Zhuang Z, Cai S, Zhao Z, Zhou L, Zhang J, Wang PH, Zhao J, Cui J. Main protease of SARS-CoV-2 serves as a bifunctional molecule in restricting type I interferon antiviral signaling. Signal Transduct Target Ther. 2020 Oct 6;5(1):221. doi: 10.1038/s41392-020-00332-2. PMID: 33024073; PMCID: PMC7537955.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 390

@@ Line 5: / Line 5: @@
 Non-structural protein 5 (nsp5) is the most highly conserved non-structural protein across the Coronaviridae family, with a molecular weight of approximately 33 kDa[1].In SARS-Cov-2, it's referred to as the main protease due to its pivotal role in the cleavage of polyproteins. Nsp5 has up to 11 conserved cleavage sites on the polyprotein and first undergoes autolytic cleavage to release itself, after which it cleaves at these sites to release other non-structural proteins.
 Nsp5 consists of three structural domains: two N-terminal domains with protease activity and a C-terminal domain composed of &#945;-helices. The active form of SARS-CoV-2 nsp5 is a homodimer that recognizes substrates approximately 10 amino acid residues in length, but exhibits selectivity at only four specific positions[2]. In addition to cleaving viral proteins, nsp5 also suppresses the host innate immune response by degrading host protein factors[3]. In this project, our primary goal is to express functional nsp5 in E.coli BL21 in preparation for in vivo inhibitor screening. Additionally, since nsp5 has the ability to recognize and cleave specific sequences, we also aim to develop it as a tool enzyme for removing recombinant tags during protein purification.
-Reference
+<b>Reference:</b><br>
+<br>
 . Jin Z, Du X, Xu Y, Deng Y, Liu M, Zhao Y, Zhang B, Li X, Zhang L, Peng C, Duan Y, Yu J, Wang L, Yang K, Liu F, Jiang R, Yang X, You T, Liu X, Yang X, Bai F, Liu H, Liu X, Guddat LW, Xu W, Xiao G, Qin C, Shi Z, Jiang H, Rao Z, Yang H. Structure of M(pro) from SARS-CoV-2 and discovery of its inhibitors. Nature. 2020 Jun;582(7811):289-293. doi: 10.1038/s41586-020-2223-y. Epub 2020 Apr 9. PMID: 32272481.
 . Zhao Y, Zhu Y, Liu X, Jin Z, Duan Y, Zhang Q, Wu C, Feng L, Du X, Zhao J, Shao M, Zhang B, Yang X, Wu L, Ji X, Guddat LW, Yang K, Rao Z, Yang H. Structural basis for replicase polyprotein cleavage and substrate specificity of main protease from SARS-CoV-2. Proc Natl Acad Sci U S A. 2022 Apr 19;119(16):e2117142119. doi: 10.1073/pnas.2117142119. Epub 2022 Apr 5. PMID: 35380892; PMCID: PMC9172370.