Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate
The prediction that the pandemic is progressing towards becoming endemic does not change the fact that COVID-19 can still be fatal for individuals with weak immune systems. Therefore, anti-COVID drugs are still needed, even when the disease becomes endemic. With regards to SARS-CoV-2, the roles of 3...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Penerbit Universiti Kebangsaan Malaysia
2023
|
| Online Access: | http://journalarticle.ukm.my/23353/1/SD%204.pdf |
| _version_ | 1796933774139719680 |
|---|---|
| author | Putra, Gede Eka Perdana Ulfah, Maria Hafizh, Naufal Prabandari, Erwahyuni Endang Firdayani, Helianti, Is |
| author_facet | Putra, Gede Eka Perdana Ulfah, Maria Hafizh, Naufal Prabandari, Erwahyuni Endang Firdayani, Helianti, Is |
| author_sort | Putra, Gede Eka Perdana |
| collection | UKM |
| description | The prediction that the pandemic is progressing towards becoming endemic does not change the fact that COVID-19 can still be fatal for individuals with weak immune systems. Therefore, anti-COVID drugs are still needed, even when the disease becomes endemic. With regards to SARS-CoV-2, the roles of 3CL protease are crucial in the formation of new virus particles. Therefore, inhibiting the function of these viral proteases will directly prevent viral replication in the human body. In this study, we report the production of a recombinant 3CL protease from SARS-CoV-2 in E. coli BL21 (DE3), which has not been extensively studied in Indonesia. The purified 3CL protease exhibited high solubility and functional activity. Additionally, the recombinant enzyme was characterised using the Rhodamine 110 fluorogenic peptide substrate. We showed that the recombinant 3CL protease was unstable in the presence of a DMSO concentration above 10%. Using the Rhodamine 110 fluorogenic peptide substrate, we found that the enzyme had a KM of 47.0 µM, Vmax of 0.41 RFU/s, and kcat/KM of 0.0088 RFU/μM2 . s while the IC50 of the GC376 was 13.35 nM. We also tested three bioactive compounds (catechin, emodin, and 1,4-naphthoquinone) using this recombinant protease as a protein target, and 1,4-naphthoquinone was the most promising bioactive compound in inhibiting the SARS-CoV-2 virus. |
| first_indexed | 2024-04-21T08:53:08Z |
| format | Article |
| id | ukm.eprints-23353 |
| institution | Universiti Kebangsaan Malaysia |
| language | English |
| last_indexed | 2024-04-21T08:53:08Z |
| publishDate | 2023 |
| publisher | Penerbit Universiti Kebangsaan Malaysia |
| record_format | dspace |
| spelling | ukm.eprints-233532024-04-17T04:36:21Z http://journalarticle.ukm.my/23353/ Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate Putra, Gede Eka Perdana Ulfah, Maria Hafizh, Naufal Prabandari, Erwahyuni Endang Firdayani, Helianti, Is The prediction that the pandemic is progressing towards becoming endemic does not change the fact that COVID-19 can still be fatal for individuals with weak immune systems. Therefore, anti-COVID drugs are still needed, even when the disease becomes endemic. With regards to SARS-CoV-2, the roles of 3CL protease are crucial in the formation of new virus particles. Therefore, inhibiting the function of these viral proteases will directly prevent viral replication in the human body. In this study, we report the production of a recombinant 3CL protease from SARS-CoV-2 in E. coli BL21 (DE3), which has not been extensively studied in Indonesia. The purified 3CL protease exhibited high solubility and functional activity. Additionally, the recombinant enzyme was characterised using the Rhodamine 110 fluorogenic peptide substrate. We showed that the recombinant 3CL protease was unstable in the presence of a DMSO concentration above 10%. Using the Rhodamine 110 fluorogenic peptide substrate, we found that the enzyme had a KM of 47.0 µM, Vmax of 0.41 RFU/s, and kcat/KM of 0.0088 RFU/μM2 . s while the IC50 of the GC376 was 13.35 nM. We also tested three bioactive compounds (catechin, emodin, and 1,4-naphthoquinone) using this recombinant protease as a protein target, and 1,4-naphthoquinone was the most promising bioactive compound in inhibiting the SARS-CoV-2 virus. Penerbit Universiti Kebangsaan Malaysia 2023 Article PeerReviewed application/pdf en http://journalarticle.ukm.my/23353/1/SD%204.pdf Putra, Gede Eka Perdana and Ulfah, Maria and Hafizh, Naufal and Prabandari, Erwahyuni Endang and Firdayani, and Helianti, Is (2023) Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate. Sains Malaysiana, 52 (12). pp. 3395-3405. ISSN 0126-6039 https://www.ukm.my/jsm/english_journals/vol52num12_2023/contentsVol52num12_2023.html |
| spellingShingle | Putra, Gede Eka Perdana Ulfah, Maria Hafizh, Naufal Prabandari, Erwahyuni Endang Firdayani, Helianti, Is Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title | Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title_full | Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title_fullStr | Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title_full_unstemmed | Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title_short | Characterisation of recombinant 3CL protease from SARS-CoV-2 produced in E. coli BL21 (DE3) for screening anti-covid drug candidates using rhodamine 110-synthetic peptide conjugate as a substrate |
| title_sort | characterisation of recombinant 3cl protease from sars cov 2 produced in e coli bl21 de3 for screening anti covid drug candidates using rhodamine 110 synthetic peptide conjugate as a substrate |
| url | http://journalarticle.ukm.my/23353/1/SD%204.pdf |
| work_keys_str_mv | AT putragedeekaperdana characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate AT ulfahmaria characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate AT hafizhnaufal characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate AT prabandarierwahyuniendang characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate AT firdayani characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate AT heliantiis characterisationofrecombinant3clproteasefromsarscov2producedinecolibl21de3forscreeninganticoviddrugcandidatesusingrhodamine110syntheticpeptideconjugateasasubstrate |