A novel genetic system for recombinant protein secretion in the Antarctic Pseudoalteromonas haloplanktis TAC125
Tóm tắt
The final aim of recombinant protein production is both to have a high specific production rate and a high product quality. It was already shown that using cold-adapted bacteria as host vectors, some "intractable" proteins can be efficiently produced at temperature as low as 4°C. A novel genetic system for the production and secretion of recombinant proteins in the Antarctic Gram-negative bacterium Pseudoalteromonas haloplanktis TAC125 was set up. This system aims at combining the low temperature recombinant product production with the advantages of extra-cellular protein targeting. The psychrophilic α-amylase from Pseudoalteromonas haloplanktis TAB23 was used as secretion carrier. Three chimerical proteins were produced by fusing intra-cellular proteins to C-terminus of the psychrophilic α-amylase and their secretion was analysed. Data reported in this paper demonstrate that all tested chimeras were translocated with a secretion yield always higher than 80%. Data presented here demonstrate that the "cold" gene-expression system is efficient since the secretion yield of tested chimeras is always above 80%. These secretion performances place the α-amylase derived secretion system amongst the best heterologous secretion systems in Gram-negative bacteria reported so far. As for the quality of the secreted passenger proteins, data presented suggest that the system also allows the correct disulphide bond formation of chimera components, secreting a fully active passenger.
Tài liệu tham khảo
Mergulhao FJ, Summers DK, Monteiro GA: Recombinant protein secretion in Escherichia coli. Biotechnol Adv. 2005, 23: 177-202. 10.1016/j.biotechadv.2004.11.003.
Speed MA, Wang DI, King J: Specific aggregation of partially folded polypeptide chains: the molecular basis of inclusion body composition. Nat Biotechnol. 1996, 14: 1283-7. 10.1038/nbt1096-1283.
Georgiou G, Valax P: Expression of correctly folded proteins in Escherichia coli. Curr Opin Biotechnol. 1996, 7: 190-7. 10.1016/S0958-1669(96)80012-7.
Baneyx F: Recombinant protein expression in Escherichia coli. Curr Opin Biotechnol. 1999, 10: 411-21. 10.1016/S0958-1669(99)00003-8.
Duilio A, Tutino ML, Marino G: Recombinant protein production in Antarctic Gram-negative bacteria. Methods Mol Biol. 2004, 267: 225-237.
Médigue C, Krin E, Pascal G, Barbe V, Bernsel A, Bertin PN, Cheung F, Cruveiller S, D'Amico S, Duilio A, Fang G, Feller G, Ho C, Mangenot S, Marino G, Nilsson J, Parrilli E, Rocha EPC, Rouy Z, Sekowska A, Tutino ML, Vallenet D, von Heijne G, Danchin A: Coping with cold: the genome of the versatile marine Antarctica bacterium Pseudoalteromonas haloplanktis TAC125. Genome Research. 2005, 15: 1325-35. 10.1101/gr.4126905.
Duilio A, Marino G, Mele A, Sannia G, Tutino ML: Sistema di espressione di proteine ricombinanti a basse temperature. 2003, Ufficio Italiano Brevetti e Marchi n. RM2003/A000155
Vigentini I, Merico A, Tutino ML, Compagno C, Marino G: Optimization of recombinant Human Nerve Growth Factor production in the psychrophilic Pseudoalteromonas haloplanktis. J Biotechnol. 2006, PMID: 16859797,
Papa R, Rippa V, Sannia G, Marino G, Duilio A: An effective cold inducible expression system developed in Pseudoalteromonas haloplanktis TAC125. J Biotechnol. 2007, 127: 199-210. 10.1016/j.jbiotec.2006.07.003.
Feller G, Lonhienne C, Deroanne C, Libioulle J, Van Beeumen J, Gerday C: Purification, characterization, and nucleotide sequence of the thermolabile alpha-amylase from the Antarctic psychrotroph Alteromonas haloplanctis A23. J Biol Chem. 1992, 267: 5217-5221.
Aghajari N, Feller G, Gerday C, Haser R: Crystal structures of the psychrophilic alpha-amylase from Alteromonas haloplanctis in its native form and complexed with an inhibitor. Protein Sci. 1998, 7: 564-572.
Feller G, D'Amico S, Benotmane AM, Joly F, Van Beeumen J, Gerday C: Characterization of the C-terminal propeptide involved in bacterial wall spanning of alpha-amylase from the psychrophile Alteromonas haloplanctis. J Biol Chem. 1998, 273: 12109-12115. 10.1074/jbc.273.20.12109.
Tutino ML, Parrilli E, Giaquinto L, Duilio A, Sannia G, Feller G, Marino G: Secretion of α-amylase from Pseudoalteromonas haloplanktis TAB23: two different pathways in different hosts. J Bacteriol. 2002, 184: 5814-7. 10.1128/JB.184.20.5814-5817.2002.
Andreotti G, Tutino ML, Sannia G, Marino G, Cubellis MV: Indole-3-glycerol-phosphate synthetase from Sulfolobus solfataricus as a model for studying thermostable TIM-barrel enzymes. Biochim Biophys Acta. 1994, 1208: 310-315.
Madonna S, Papa R, Birolo L, Autore F, Doti N, Marino G, Quemeneur E, Sannia G, Tutino ML, Duilio A: The thiol-disulphide oxidoreductase system in the cold-adapted bacterium Pseudoalteromonas haloplanktis TAC 125: discovery of a novel disulfide oxidoreductase enzyme. Extremophiles. 2006, 10: 41-51. 10.1007/s00792-005-0470-3.
Holmgren A: Thioredoxin catalyzes the reduction of insulin disulfides by dithiothreitol and dihydrolipoamide. J Biol Chem. 1979, 254: T9113-9119.
Georgiou G, Segatori L: Preparative expression of secreted proteins in bacteria: status report and future prospects. Curr Opin Biotechnol. 2005, 16: 538-45. 10.1016/j.copbio.2005.07.008.
Cusano AM, Parrilli E, Duilio A, Sannia G, Marino G, Tutino ML: Secretion of psychrophilic alpha-amylase deletion mutants in Pseudoalteromonas haloplanktis TAC125. FEMS Microbiol Lett. 2006, 258: 67-71. 10.1111/j.1574-6968.2006.00193.x.
Eom GT, Song JK, Ahn JH, Seo YS, Rhee JS: Enhancement of the efficiency of secretion of heterologous lipase in Escherichia coli by directed evolution of the ABC transporter system. Appl Environ Microbiol. 2005, 71: 3468-74. 10.1128/AEM.71.7.3468-3474.2005.
Sugamata Y, Shiba T: Improved secretory production of recombinant proteins by random mutagenesis of hlyB, an alpha-hemolysin transporter from Escherichia coli. Appl Environ Microbiol. 2005, 71: 656-62. 10.1128/AEM.71.2.656-662.2005.
Zhang G, Brokx S, Weiner JH: Extracellular accumulation of recombinant proteins fused to the carrier protein YebF in Escherichia coli. Nat Biotechnol. 2006, 24: 100-4. 10.1038/nbt1174.
Hennig M, Darimont B, Sterner R, Kirschner K, Jansonius JN: 2.0 A structure of indole-3-glycerol phosphate synthase from the hyperthermophile Sulfolobus solfataricus : possible determinants of protein stability. Structure. 1995, 3: 1295-306. 10.1016/S0969-2126(01)00267-2.
Schneider B, Knochel T, Darimont B, Hennig M, Dietrich S, Babinger K, Kirschner K, Sterner R: Role of the N-terminal extension of the (beta alpha)8-barrel enzyme indole-3-glycerol phosphate synthase for its fold, stability, and catalytic activity. Biochemistry. 2005, 44: 16405-12. 10.1021/bi051640n.
Duilio A, Madonna S, Tutino ML, Pirozzi M, Sannia G, Marino G: Promoters from a cold-adapted bacterium: definition of a consensus motif and molecular characterization of UP regulative elements. Extremophiles. 2004, 8: 125-32. 10.1007/s00792-003-0371-2.
Hanahan D: Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983, 166: 557-80. 10.1016/S0022-2836(83)80284-8.
Sambrook J, Russell DW: Molecular Cloning. A Laboratory Manual. Edited by: Spring Harbor Laboratory, Cold Spring Harbor NY. 2001, 3
Jones JV, Mansour M, James H, Sadi D, Carr RI: A substrate amplification system for enzyme-linked immunoassays. II. Demonstration of its applicability for measuring anti-DNA antibodies. J Immunol Methods. 1989, 118: 79-84.
O'Callaghan CH, Morris A, Kirby SM, Shingler AH: Novel method for detection of β-lactamase by using a chromogenic cephalosporin substrate. Antimicrob Ag Chemother. 1972, 1: 283-288.