Evaluation of induction conditions for plasmid pQE-30 stability and 503 antigen of Leishmania i. chagasi expression in E. coli M15

被引：20
作者：
Ribeiro, Vitor Troccoli [1 ]
Asevedo, Estefani Alves [1 ]
Costa de Paiva Vasconcelos, Luan Tales [1 ]
Oliveira Filho, Marcos Antonio [1 ]
de Araujo, Jaciara Silva [1 ]
de Macedo, Gorete Ribeiro [1 ]
de Sousa Junior, Francisco Caninde [2 ]
dos Santos, Everaldo Silvino [1 ]
机构：
[1] Univ Fed Rio Grande do Norte, Chem Engn Dept, Ave Sen Salgado Filho,3000, BR-59078970 Natal, RN, Brazil
[2] Univ Fed Rio Grande do Norte, Pharm Dept, BR-59012570 Natal, RN, Brazil
来源：
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY | 2019年 / 103卷 / 16期
关键词：
Leishmania i; chagasi; Induction; 503; antigen; Plasmid stability; ACETIC-ACID ACCUMULATION; ESCHERICHIA-COLI; CELL-GROWTH; RECOMBINANT PROTEINS; IPTG CONCENTRATION; INFANTUM-CHAGASI; OPTIMIZATION; PURIFICATION; TEMPERATURE; CLONING;
D O I：
10.1007/s00253-019-09948-z
中图分类号：
Q81 [生物工程学（生物技术）]; Q93 [微生物学];
学科分类号：
071005 ; 0836 ; 090102 ; 100705 ;
摘要：
The present study aimed to evaluate the influence of induction conditions (IPTG concentration, temperature, and induction time) on the plasmid pQE-30 stability and 503 antigen expression of Leishmania i. chagasi in Escherichia coli M15. Batch cultures were performed at 37 degrees C and induced by the addition of different IPTG concentrations (0.01 to 1.5 mM). Subsequently, experiments were carried out at different temperatures (27 to 42 degrees C), evaluating the influence of induction time (0.5 to 6 h after the start of the culture). The results showed that IPTG toxicity caused a metabolic stress in the cells and, consequently, the microorganism growth reduced. The induction with IPTG may also be associated with the plasmid pQE-30 instability, due to metabolic burden imposed by the recombinant protein expression. The optimal conditions for 503 antigen expression of Leishmania i. chagasi in Escherichia coli M15 were an IPTG concentration of 1.0 mM, temperature of 37 degrees C, and induction time of 2 h. The maximum antigen concentration obtained was 0.119 +/- 0.009 g/L, about seven times higher than the lowest concentration. Therefore, the results showed that 503 antigen can be produced in laboratory; however, it requires more studies to minimize the plasmid instability and improve to industrial scale.
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收藏
页码：6495 / 6504
页数：10
相关论文
共 38 条
[1]  
Afonso Margarida S, 2015, Biotechnol Rep (Amst), V5, P7, DOI 10.1016/j.btre.2014.10.008
[2]   Overcoming challenges for amplified expression of recombinant proteins using Escherichia coli [J].
Ahmad, Irshad ;
Nawaz, Nighat ;
Darwesh, Nizam M. ;
Rahman, Sadeeq Ur ;
Mustafa, Mohammad Z. ;
Khan, Sher B. ;
Patching, Simon G. .
PROTEIN EXPRESSION AND PURIFICATION, 2018, 144 :12-18
[3]   Identifying conditions for inducible protein production in E. coli:: combining a fed-batch and multiple induction approach [J].
Aucoin, Marc G. ;
McMurray-Beaulieu, Virginie ;
Poulin, Frederic ;
Boivin, Eric B. ;
Chen, Jingkui ;
Ardelean, Francisc M. ;
Cloutier, Mathieu ;
Choi, Young J. ;
Miguez, Carlos B. ;
Jolicoeur, Mario .
MICROBIAL CELL FACTORIES, 2006, 5 (1)
[4]   Effect of low temperature on stability of θ-type plasmids in Carnobacterium maltaromaticum [J].
Bohaychuk, Valerie M. ;
Van Belkum, Marco J. ;
Stiles, Michael E. ;
McMullen, Lynn M. .
FEMS MICROBIOLOGY LETTERS, 2008, 280 (01) :14-20
[5]   Silver staining of proteins in polyacrylamide gels [J].
Chevallet, Mireille ;
Luche, Sylvie ;
Rabilloud, Thierry .
NATURE PROTOCOLS, 2006, 1 (04) :1852-1858
[6]   Batch production of a silk-elastin-like protein in E. coli BL21 (DE3): key parameters for optimisation [J].
Collins, Tony ;
Azevedo-Silva, Joao ;
da Costa, Andre ;
Branca, Fernando ;
Machado, Raul ;
Casal, Margarida .
MICROBIAL CELL FACTORIES, 2013, 12
[7]   SYNTHESIS, CHARACTERIZATION AND INTERACTION STUDIES OF AN ANALOG OF CcdA ANTITOXIN BY STEADY STATE FLUORESCENCE. [J].
Cotrim, Camila Aparecida ;
Garrido, Saulo Santesso ;
Trovatti, Eliane ;
Marchetto, Reinaldo .
QUIMICA NOVA, 2010, 33 (04) :841-845
[8]   Cloning, Auto-induction Expression, and Purification of rSpaA Swine Erysipelas Antigen [J].
da Silva, Adilson Jose ;
da Costa Iemma, Monica Rosas ;
Luperni Horta, Antonio Carlos ;
Sargo, Cintia Regina ;
Camargo Giordano, Raquel de Lima ;
Giordano, Roberto de Campos ;
Zangirolami, Teresa Cristina ;
Marques Novo, Maria Teresa .
CURRENT MICROBIOLOGY, 2012, 65 (04) :369-374
[9]   Recovery and purification of recombinant 503 antigen of Leishmania infantum chagasi using expanded bed adsorption chromatography [J].
de Sousa Junior, Francisco Caninde ;
Ferreira Vaz, Michelle Rossana ;
de Araujo Padilha, Carlos Eduardo ;
Chiberio, Abimaelle Silva ;
Arantes Martins, Daniella Regina ;
de Macedo, Gorete Ribeiro ;
dos Santos, Everaldo Silvino .
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 2015, 986 :1-7
[10]   Cloning and expression of protease ClpP from Streptococcus pneumoniae in Escherichia coli: Study of the influence of kanamycin and IPTG concentration on cell growth, recombinant protein production and plasmid stability [J].
Einsfeldt, Karen ;
Severo Junior, Joao Baptista ;
Correa Argondizzo, Ana Paula ;
Medeiros, Marco Alberto ;
Moitinho Alves, Tito Livio ;
Almeida, Rodrigo Volcan ;
Larentis, Ariane Leites .
VACCINE, 2011, 29 (41) :7136-7143
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