The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts
The thermolabile haemolysin (tlh) of Vibrio parahaemolyticus (Vptlh) from V. parahaemolyticus is a multiple-function enzyme, initially describes as a haemolytic factor activated by lecithin and phospholipase A2 enzymatic activity (Shinoda, 1991; Vazquez-Morado, 2021; Yanagase et al., 1970). Until no...
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The Korean Society of Fisheries and Aquatic Science
2023-03-01
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| Series: | Fisheries and Aquatic Sciences |
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| Online Access: | http://www.e-fas.org/archive/view_article?doi=10.47853/FAS.2023.e17 |
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| author | Tran Thi Huyen Ha Phuong Trang Nguyen Thi-Ngan Bui Dinh-Thanh Le Pham Tan Quoc Trinh Ngoc Nam |
| author_facet | Tran Thi Huyen Ha Phuong Trang Nguyen Thi-Ngan Bui Dinh-Thanh Le Pham Tan Quoc Trinh Ngoc Nam |
| author_sort | Tran Thi Huyen |
| collection | DOAJ |
| description | The thermolabile haemolysin (tlh) of Vibrio parahaemolyticus (Vptlh) from V. parahaemolyticus is a multiple-function enzyme, initially describes as a haemolytic factor activated by lecithin and phospholipase A2 enzymatic activity (Shinoda, 1991; Vazquez-Morado, 2021; Yanagase et al., 1970). Until now, the tlh structure has hypothesized including N-terminal and C-terminal domain, but what domain of the Vptlh structure does the haemolytic activity has not been refined yet. In this study, a 450-bp VpTLH nucleotide sequence of the entire Vptlh gene encoded the C-terminal domain cloned firstly to examine its responsibility in the activity of the Vptlh. The C-terminal domain fused with a 6-His-tag named the His-tag-VpC-terminal domain was expressed successfully in soluble form in the BL21 (DE3) PlysS cell. Remarkably, both expression and purification results confirmed a high agreement in the molecular weight of the His-tag-VpC-terminal domain was 47 kDa. This work showed the His-tag-VpC-terminal domain lysed the erythrocyte membranes in the blood agar and the phosphate buffered saline (0.9%) media without adding the lecithin substrate of the phospholipase enzyme. Haemolysis occurred at all tested diluted concentrations of His-tag-VpC-terminal domain (p < 0.05), providing evidence for the independent haemolytic activity of the His-tag-VpC-terminal domain. The content of 100 μg of the His-tag-VpC-terminal domain brought the highest haemolytic activity of 80% compared to that in the three remaining contents. Significantly, the His-tag-VpC-terminal domain demonstrated not to involve the phospholipase activity in Luria-Bertani agar supplemented with 1% (vol/vol) egg yolk emulsion. All results proved the vital responsibility of the His-tag-VpC-terminal domain in causing the haemolytic activity without the required activation by the phospholipase enzyme. Raw extracts of Phellinus igniarus and Phellinus pipi at 10-1 mg/mL inhibited the haemolytic activity of the His-tag-VpC-terminal domain from 67.7% to 87.42%, respectively. Hence applying the His-tag-VpC-terminal domain as a simple biological material to evaluate quickly potential derivatives against the Vptlh in vivo conditions will accessible and more advantageous than using the whole of the Vptlh. |
| first_indexed | 2024-04-09T19:40:24Z |
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| id | doaj.art-cf11fea4e59044be896b7587d8699f05 |
| institution | Directory Open Access Journal |
| issn | 2234-1757 |
| language | English |
| last_indexed | 2024-04-09T19:40:24Z |
| publishDate | 2023-03-01 |
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| series | Fisheries and Aquatic Sciences |
| spelling | doaj.art-cf11fea4e59044be896b7587d8699f052023-04-04T08:47:54ZengThe Korean Society of Fisheries and Aquatic ScienceFisheries and Aquatic Sciences2234-17572023-03-0126320421510.47853/FAS.2023.e17The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extractsTran Thi Huyen0Ha Phuong Trang1Nguyen Thi-Ngan2Bui Dinh-Thanh3Le Pham Tan Quoc4Trinh Ngoc Nam5Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamInstitute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamInstitute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamInstitute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamInstitute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamInstitute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City, Ho Chi Minh 71408, VietnamThe thermolabile haemolysin (tlh) of Vibrio parahaemolyticus (Vptlh) from V. parahaemolyticus is a multiple-function enzyme, initially describes as a haemolytic factor activated by lecithin and phospholipase A2 enzymatic activity (Shinoda, 1991; Vazquez-Morado, 2021; Yanagase et al., 1970). Until now, the tlh structure has hypothesized including N-terminal and C-terminal domain, but what domain of the Vptlh structure does the haemolytic activity has not been refined yet. In this study, a 450-bp VpTLH nucleotide sequence of the entire Vptlh gene encoded the C-terminal domain cloned firstly to examine its responsibility in the activity of the Vptlh. The C-terminal domain fused with a 6-His-tag named the His-tag-VpC-terminal domain was expressed successfully in soluble form in the BL21 (DE3) PlysS cell. Remarkably, both expression and purification results confirmed a high agreement in the molecular weight of the His-tag-VpC-terminal domain was 47 kDa. This work showed the His-tag-VpC-terminal domain lysed the erythrocyte membranes in the blood agar and the phosphate buffered saline (0.9%) media without adding the lecithin substrate of the phospholipase enzyme. Haemolysis occurred at all tested diluted concentrations of His-tag-VpC-terminal domain (p < 0.05), providing evidence for the independent haemolytic activity of the His-tag-VpC-terminal domain. The content of 100 μg of the His-tag-VpC-terminal domain brought the highest haemolytic activity of 80% compared to that in the three remaining contents. Significantly, the His-tag-VpC-terminal domain demonstrated not to involve the phospholipase activity in Luria-Bertani agar supplemented with 1% (vol/vol) egg yolk emulsion. All results proved the vital responsibility of the His-tag-VpC-terminal domain in causing the haemolytic activity without the required activation by the phospholipase enzyme. Raw extracts of Phellinus igniarus and Phellinus pipi at 10-1 mg/mL inhibited the haemolytic activity of the His-tag-VpC-terminal domain from 67.7% to 87.42%, respectively. Hence applying the His-tag-VpC-terminal domain as a simple biological material to evaluate quickly potential derivatives against the Vptlh in vivo conditions will accessible and more advantageous than using the whole of the Vptlh. http://www.e-fas.org/archive/view_article?doi=10.47853/FAS.2023.e17A 450-bp VpTLH nucleotide sequencethermolabile haemolysin gene (tlh)treatment haemolytic activity by Phellinus spVibrio parahaemolyticusC-terminal domain |
| spellingShingle | Tran Thi Huyen Ha Phuong Trang Nguyen Thi-Ngan Bui Dinh-Thanh Le Pham Tan Quoc Trinh Ngoc Nam The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts Fisheries and Aquatic Sciences A 450-bp VpTLH nucleotide sequence thermolabile haemolysin gene (tlh) treatment haemolytic activity by Phellinus sp Vibrio parahaemolyticus C-terminal domain |
| title | The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts |
| title_full | The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts |
| title_fullStr | The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts |
| title_full_unstemmed | The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts |
| title_short | The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts |
| title_sort | responsibility of c terminal domain in the thermolabile haemolysin activity of vibrio parahaemolyticus and inhibition treatments by phellinus sp extracts |
| topic | A 450-bp VpTLH nucleotide sequence thermolabile haemolysin gene (tlh) treatment haemolytic activity by Phellinus sp Vibrio parahaemolyticus C-terminal domain |
| url | http://www.e-fas.org/archive/view_article?doi=10.47853/FAS.2023.e17 |
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