Molecular analysis of hemolysin-mediated secretion of a human interleukin-6 fusion protein in Salmonella typhimurium |
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| Authors: | Li Y Hess C von Specht B Hahn H P |
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| Affiliation: | 1. Univ. Bourgogne Franche-Comté, AgroSup Dijon, PAM UMR A 02.102, F-21000 Dijon, France;2. Welience (SATT Grand-Est), Hall de Technologie Alimentaire, F-21000 Dijon, France;1. Department of Pharmaceutics, School of Pharmacy, Nanjing Medical University, Nanjing, 211116, China;2. Department of Dermatology, Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School, Nanjing, 210093, China;3. Department of Oncology, The Affiliated Hospital of Kangda College of Nanjing Medical University & the First People’s Hospital of Lianyungang, Lianyungang, 222000, China;1. State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China;2. Key laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi 214122, China;3. Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, China;1. School of Molecular Cell Biology & Biotechnology, Tel Aviv University, Ramat Aviv, Israel;2. Department of Biology, Stanford University, Stanford, CA 94305-5020, USA;3. School of Molecular Biology and Ecology of Plants, Tel Aviv University, Ramat Aviv, Israel;1. Department of Life Science, College of Natural Science, Chung-Ang University, Seoul 156-756, South Korea;2. College of Pharmacy, Chung-Ang University, Seoul 156-756, South Korea;3. Pathology Division, National Institute of Fisheries Science (NIFS), Busan 46083, South Korea;4. Green Cross Veterinary Products Co. LTD., Yongin, Kyunggi-Do, South Korea;5. Bio-Integration Research Center for Nutra-Pharmaceutical Epigenetics, Chung-Ang University, Seoul 156-756, South Korea |
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| Abstract: | Previously, we reported a plasmid-bearing Salmonella typhimurium strain capable of secreting human interleukin-6 (hIL-6) when genetically fused to the Escherichia coli hemolysin transport signal (HlyA(S)). Stationary phase culture supernatants of this strain revealed three major forms of hIL-6-HlyA(S) fusion protein (apparent molecular masses 32.4, 30.3, 27.0 kDa), at which the largest protein presumably represented full-length hIL-6-HlyA(S). The biological activity of the hIL-6-HlyA(S) protein mixture was similar to that of mature hIL-6. Accumulation of hIL-6-HlyA(S) in the culture supernatant occurred only during the initial growth phase, whereas in stationary phase and under in vitro conditions successive cleavage into the two truncated forms was observed. On the other hand, in whole cell lysates only full-length hIL-6-HlyA(S) could be detected, accounting for more than 50% of the totally synthesized protein. Upon cell fractionation, cellular hIL-6-HlyA(S) was exclusively found in the membrane fraction. These results suggest, that in S. typhimurium production and secretion of hIL-6-HlyA(S) is restricted to growing cells. A specific processing by a Salmonella-derived protease did not affect the biological activity of the fusion protein. |
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| Keywords: | Fusion protein Hemolysin Interleukin-6 Protein secretion Proteolytic cleavage |
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