Fisheries and Aquatic Sciences

한국수산과학회 (The Korean Society of Fisheries and Aquatic Science)
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The responsibility of C-terminal domain in the thermolabile haemolysin activity of Vibrio parahaemolyticus and inhibition treatments by Phellinus sp. extracts

  • Tran Thi Huyen (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City) ;
  • Ha Phuong Trang (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City) ;
  • Nguyen Thi-Ngan (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City) ;
  • Bui Dinh-Thanh (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City) ;
  • Le Pham Tan Quoc (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City) ;
  • Trinh Ngoc Nam (Institute of Biotechnology and Food Technology, Industrial University of Ho Chi Minh City)
  • 투고 : 2022.10.07
  • 심사 : 2023.01.19
  • 발행 : 2023.03.31
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초록

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 ㎍ 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.

키워드

과제정보

Tran Thi Huyen conceived and designed the experiments, performed the experiments, analyzed the data, prepared Figs and/ or Tables, authored or reviewed drafts of the paper and approved the final draft. Phuong-Trang Ha, Bui Dinh Thanh performed the experiments, analyzed the data, prepared Figs and/or Tables. Nguyen Thi Ngan conceived and designed the extracts experiments from phellinus sp. Trinh Ngoc Nam, Le Pham Tan Quoc critically reviewed the manuscript and approved the final draft. All authors read and approved the final manuscript.

참고문헌

  1. Bej AK, Patterson DP, Brasher CW, Vickery MCL, Jones DD, Kaysner CA. Detection of total and hemolysin-producing Vibrio parahaemolyticus in shellfish using multiplex PCR amplification of tl, tdh and trh. J Microbiol Methods. 1999;36:215-25. https://doi.org/10.1016/S0167-7012(99)00037-8
  2. Chow MKM, Amin AA, Fulton KF, Fernando T, Kamau L, Batty C, et al. The REFOLD database: a tool for the optimization of protein expression and refolding. Nucleic Acids Res. 2006;34:D207-12. https://doi.org/10.1093/nar/gkj080
  3. Food and Agriculture Organization [FAO], World Health Organization [WHO]. Risk assessment tools for Vibrio parahaemolyticus and Vibrio vulnificus associated with seafood: meeting report. Rome: FAO; 2020.
  4. Fujino T, Okuno Y, Nakada D, Aoyama A, Fukai K, Mukai T, et al. On the bacteriological examination of Shirasu food poisoning. Med J Osaka Univ. 1953;4:299-304.
  5. Guerrero A, Lizarraga-Partida ML, Gil Rodriguez BG, Licea-Navarro AF, Revilla-Castellanos VJ, Wong-Chang I, et al. Genetic analysis of Vibrio parahaemolyticus O3:K6 strains that have been isolated in Mexico since 1998. PLOS ONE. 2017;12:e0169722.
  6. He P, Zhang Y, Li N. The phytochemistry and pharmacology of medicinal fungi of the genus Phellinus: a review. Food Funct. 2021;12:1856-81. https://doi.org/10.1039/D0FO02342F
  7. Jang KK, Lee ZW, Kim B, Jung YH, Han HJ, Kim MH, et al. Identification and characterization of Vibrio vulnificus plpA encoding a phospholipase A2 essential for pathogenesis. J Biol Chem. 2017;292:17129-43. https://doi.org/10.1074/jbc.M117.791657
  8. Jia A, Woo NYS, Zhang XH. Expression, purification, and characterization of thermolabile hemolysin (TLH) from Vibrio alginolyticus. Dis Aquat Organ. 2010;90:121-7. https://doi.org/10.3354/dao02225
  9. Li L, Mou X, Nelson DR. Characterization of Plp, a phosphatidylcholine-specific phospholipase and hemolysin of Vibrio anguillarum. BMC Microbiol. 2013;13:271.
  10. Mahmuda ZH, Kassu A, Mohammada A, Yamatoa M, Bhuiyanb NA, Balakrish Nairb G, et al. Isolation and molecular characterization of toxigenic Vibrio parahaemolyticus from the Kii Channel, Japan. Microbiol Res. 2006;161:25-37. https://doi.org/10.1016/j.micres.2005.04.005
  11. Mazzarino L, Loch-Neckel G, Bubniak LS, Ourique F, Otsuka I, Halil S, et al. Nanoparticles made from xyloglucan-block-polycaprolactone copolymers: safety assessment for drug delivery. Toxicol Sci. 2015;147:104-15. https://doi.org/10.1093/toxsci/kfv114
  12. McCarthy SA, DePaola A, Cook DW, Kaysner CA, Hill WE. Evaluation of alkaline phosphatase- and digoxigenin-labelled probes for detection of the thermolabile hemolysin (tlh) gene of Vibrio parahaemolyticus. Lett Appl Microbiol. 1999;28:66-70. https://doi.org/10.1046/j.1365-2672.1999.00467.x
  13. Meparambu Prabhakaran D, Ramamurthy T, Thomas S. Genetic and virulence characterisation of Vibrio parahaemolyticus isolated from Indian coast. BMC Microbiol. 2020;20:62.
  14. Nghia NT, Oanh DTH, Phu TQ, Tuan PA. Isolation and determination of the ability to cause acute hepatopancreatic necrosis syndrome of Vibrio parahaemolitycus bacteria isolated from cultured shrimp in Bac Lieu province. Tap chi Khoa hoc Truong Dai hoc Can Tho. 2015;39:99-107.
  15. Shinoda S. Protein toxins produced by pathogenic vibrios. J Nat Toxins. 1999;8:259-69.
  16. Shinoda S, Matsuoka H, Tsuchie T, Miyoshi SI, Yamamoto S, Taniguchi H, et al. Purification and characterization of a lecithin-dependent haemolysin from Escherichia coli transformed by a Vibrio parahaemolyticus gene. J Gen Microbiol. 1991;137:2705-11. https://doi.org/10.1099/00221287-137-12-2705
  17. Singh SM, Panda AK. Solubilization and refolding of bacterial inclusion body proteins. J Biosci Bioeng. 2005;99:303-10. https://doi.org/10.1263/jbb.99.303
  18. Su YC, Liu C. Vibrio parahaemolyticus: a concern of seafood safety. Food Microbiol. 2007;24:549-58. https://doi.org/10.1016/j.fm.2007.01.005
  19. Sun B, Zhang XH, Tang X, Wang S, Zhong Y, Chen J, et al. A single residue change in Vibrio harveyi hemolysin results in the loss of phospholipase and hemolytic activities and pathogenicity for turbot (Scophthalmus maximus). J Bacteriol. 2007;189:2575-9. https://doi.org/10.1128/JB.01650-06
  20. Taniguchi H, Hirano H, Kubomura S, Higashi K, Mizuguchi Y. Comparison of the nucleotide sequences of the genes for the thermostable direct hemolysin and the thermolabile hemolysin from Vibrio parahaemolyticus. Microb Pathog. 1986;1:425-32. https://doi.org/10.1016/0882-4010(86)90004-5
  21. Thanh NT, Tuan NN, Kuo PC, Dung DM, Phuong DL, Truong Giang DT, et al. Chemical constituents from the fruiting bodies of Phellinus igniarius. Nat Prod Res. 2018;32:2392-7. https://doi.org/10.1080/14786419.2017.1413572
  22. Tran L, Nunan L, Redman RM, Mohney LL, Pantoja CR, Fitzsimmons K, et al. Determination of the infectious nature of the agent of acute hepatopancreatic necrosis syndrome affecting penaeid shrimp. Dis Aquat Organ. 2013;105:45-55. https://doi.org/10.3354/dao02621
  23. Vazquez-Morado LE, Robles-Zepeda RE, Ochoa-Leyva A, ArvizuFlores AA, Garibay-Escobar A, Castillo-Yanez F, et al. Biochemical characterization and inhibition of thermolabile hemolysin from Vibrio parahaemolyticus by phenolic compounds. PeerJ. 2021;9:e10506.
  24. Vugia D, Cronquist A, Cartter M, Tobin-D'Angelo M, Blythe D, Smith K, et al. Preliminary FoodNet data on the incidence of infection with pathogens transmitted commonly through food---10 states, 2008. Morb Mortal Wkly Rep. 2009;58:333-7.
  25. Wang JJ, Liu KS, Sung KC, Tsai CY, Fang JY. Lipid nanoparticles with different oil/fatty ester ratios as carriers of buprenorphine and its prodrugs for injection. Eur J Pharm Sci. 2009;38:138-46. https://doi.org/10.1016/j.ejps.2009.06.008
  26. Wang R, Fang S, Wu D, Lian J, Fan J, Zhang Y, et al. Screening for a single-chain variable-fragment antibody that can effectively neutralize the cytotoxicity of the Vibrio parahaemolyticus thermolabile hemolysin. Appl Environ Microbiol. 2012;78:4967-75. https://doi.org/10.1128/AEM.00435-12
  27. Wang R, Zhong Y, Gu X, Yuan J, Saeed AF, Wang S. The pathogenesis, detection, and prevention of Vibrio parahaemolyticus. Front Microbiol. 2015;6:144.
  28. Xie ZY, Hu CQ, Chen C, Zhang LP, Ren CH. Investigation of seven Vibrio virulence genes among Vibrio alginolyticus and Vibrio parahaemolyticus strains from the coastal mariculture systems in Guangdong, China. Lett Appl Microbiol. 2005;41:202-7. https://doi.org/10.1111/j.1472-765X.2005.01688.x
  29. Xu F, Ilyas S, Hall JA, Jones SH, Cooper VS, Whistler CA. Genetic characterization of clinical and environmental Vibrio parahaemolyticus from the Northeast USA reveals emerging resident and non-indigenous pathogen lineages. Front Microbiol. 2015;6:272.
  30. Yanagase Y, Inoue K, Ozaki M, Ochi T, Amano T. Hemolysins and related enzymes of Vibrio parahaemolyticus. I. Identification and partial purification of enzymes. Biken J. 1970;13:77-92.
  31. Yanez R, Bastias R, Higuera G, Salgado O, Katharios P, Romero J, et al. Amplification of tlh gene in other Vibrionaceae specie by specie-specific multiplex PCR of Vibrio parahaemolyticus. Electron J Biotechnol. 2015;18:459-63. https://doi.org/10.1016/j.ejbt.2015.09.007
  32. Yang Y, Ye LB, Zhang JS, Liu YF, Tang QJ. Structural analysis of a bioactive polysaccharide, PISP1, from the medicinal mushroom Phellinus igniarius. Biosci Biotechnol Biochem. 2009;73:134-9. https://doi.org/10.1271/bbb.80546
  33. Yamazaki W, Ishibashi M, Kawahara R, Inoue K. Development of a loop-mediated isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus. BMC Microbiol. 2008;8:163.
  34. Zhang J, Chen XG, Li YY, Liu CS. Self-assembled nanoparticles based on hydrophobically modified chitosan as carriers for doxorubicin. Nanomed Nanotechnol Biol Med. 2007;3:258-65. https://doi.org/10.1016/j.nano.2007.08.002
  35. Zhang XH, Meaden PG, Austin B. Duplication of hemolysin genes in a virulent isolate of Vibrio harveyi. Appl Environ Microbiol. 2001;67:3161-7. https://doi.org/10.1128/AEM.67.7.3161-3167.2001
  36. Zhao Y, Tang X, Zhan W. Cloning, expressing, and hemolysis of tdh, trh and tlh genes of Vibrio parahaemolyticus. J Ocean Univ China. 2011;10:275-9.  https://doi.org/10.1007/s11802-011-1801-x