Nasira Rafi, Rabia Abbas, Nadeem Ahmed, Saad Tahir, Muhammad Akram, Mohsin Ahmad Khan, Nao Akusa Fujimura, Mariam Shafique, Kausar Malik
{"title":"玉米淀粉液作为大肠杆菌生产 rhIFN-β-1b 蛋白的自动诱导培养基","authors":"Nasira Rafi, Rabia Abbas, Nadeem Ahmed, Saad Tahir, Muhammad Akram, Mohsin Ahmad Khan, Nao Akusa Fujimura, Mariam Shafique, Kausar Malik","doi":"10.1007/s40995-024-01684-y","DOIUrl":null,"url":null,"abstract":"<div><p>Therapeutic proteins need to be produced in large quantities as they continuously gain higher shares in pharmaceutical market. Human beta interferon (hIFN-β) is used against different diseases due to its antiviral, antiproliferative and immunomodulatory nature. Considering its clinical applications, development of a simple, cost-effective and easily scalable production system to produce rhIFN-β-1b protein in high yield and quality is needed. Attempts to develop improved production methods are ongoing. Majority of previous reports propose isopropyl-β-D-1-thiogalactopyranoside (IPTG), for rhIFN-β-1b expression in <i>Escherichia coli</i>, which is costly and nonmetabolizable. Thus, in current study, rhIFN-β-1b expression was optimized in an inexpensive auto-induction medium. Corn steep liquor (CSL) was used and optimized at different concentrations. Further, the medium was also optimized for carbon sources using various glycerol concentrations. Finally, a 10 L fermentation batch was done (10% CSL and 2.5% glycerol) that produced 65 g of wet biomass and 15 g of inclusion body (wet). The purification scheme was optimized by one-step ion-exchange chromatography (IEX) for > 99% purity of the target protein. The biological activity of the required protein was assessed through cytopathic inhibition effect of virus vesicular stomatitis (VSV) on Madin-Darby bovine kidney (MDBK) cells. Moreover, cytotoxicity of the protein was also studied on different breast cancer cell lines; HCC- 1954, MCF- 7 and MDA- MB- 231. This is the first report on utilizing CSL as auto-induction media for expression and purification of rhIFN-β-1b. Thus, the suggested method can be employed for simple as well as cost-effective production of rhIFN-β-1b and other recombinant proteins.</p></div>","PeriodicalId":600,"journal":{"name":"Iranian Journal of Science and Technology, Transactions A: Science","volume":"48 5","pages":"1087 - 1098"},"PeriodicalIF":1.4000,"publicationDate":"2024-07-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Corn Steep Liquor as an Auto-Induction Medium for the Production of rhIFN-β-1b Protein in Escherichia coli\",\"authors\":\"Nasira Rafi, Rabia Abbas, Nadeem Ahmed, Saad Tahir, Muhammad Akram, Mohsin Ahmad Khan, Nao Akusa Fujimura, Mariam Shafique, Kausar Malik\",\"doi\":\"10.1007/s40995-024-01684-y\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Therapeutic proteins need to be produced in large quantities as they continuously gain higher shares in pharmaceutical market. Human beta interferon (hIFN-β) is used against different diseases due to its antiviral, antiproliferative and immunomodulatory nature. Considering its clinical applications, development of a simple, cost-effective and easily scalable production system to produce rhIFN-β-1b protein in high yield and quality is needed. Attempts to develop improved production methods are ongoing. Majority of previous reports propose isopropyl-β-D-1-thiogalactopyranoside (IPTG), for rhIFN-β-1b expression in <i>Escherichia coli</i>, which is costly and nonmetabolizable. Thus, in current study, rhIFN-β-1b expression was optimized in an inexpensive auto-induction medium. Corn steep liquor (CSL) was used and optimized at different concentrations. Further, the medium was also optimized for carbon sources using various glycerol concentrations. Finally, a 10 L fermentation batch was done (10% CSL and 2.5% glycerol) that produced 65 g of wet biomass and 15 g of inclusion body (wet). The purification scheme was optimized by one-step ion-exchange chromatography (IEX) for > 99% purity of the target protein. The biological activity of the required protein was assessed through cytopathic inhibition effect of virus vesicular stomatitis (VSV) on Madin-Darby bovine kidney (MDBK) cells. Moreover, cytotoxicity of the protein was also studied on different breast cancer cell lines; HCC- 1954, MCF- 7 and MDA- MB- 231. This is the first report on utilizing CSL as auto-induction media for expression and purification of rhIFN-β-1b. 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Corn Steep Liquor as an Auto-Induction Medium for the Production of rhIFN-β-1b Protein in Escherichia coli
Therapeutic proteins need to be produced in large quantities as they continuously gain higher shares in pharmaceutical market. Human beta interferon (hIFN-β) is used against different diseases due to its antiviral, antiproliferative and immunomodulatory nature. Considering its clinical applications, development of a simple, cost-effective and easily scalable production system to produce rhIFN-β-1b protein in high yield and quality is needed. Attempts to develop improved production methods are ongoing. Majority of previous reports propose isopropyl-β-D-1-thiogalactopyranoside (IPTG), for rhIFN-β-1b expression in Escherichia coli, which is costly and nonmetabolizable. Thus, in current study, rhIFN-β-1b expression was optimized in an inexpensive auto-induction medium. Corn steep liquor (CSL) was used and optimized at different concentrations. Further, the medium was also optimized for carbon sources using various glycerol concentrations. Finally, a 10 L fermentation batch was done (10% CSL and 2.5% glycerol) that produced 65 g of wet biomass and 15 g of inclusion body (wet). The purification scheme was optimized by one-step ion-exchange chromatography (IEX) for > 99% purity of the target protein. The biological activity of the required protein was assessed through cytopathic inhibition effect of virus vesicular stomatitis (VSV) on Madin-Darby bovine kidney (MDBK) cells. Moreover, cytotoxicity of the protein was also studied on different breast cancer cell lines; HCC- 1954, MCF- 7 and MDA- MB- 231. This is the first report on utilizing CSL as auto-induction media for expression and purification of rhIFN-β-1b. Thus, the suggested method can be employed for simple as well as cost-effective production of rhIFN-β-1b and other recombinant proteins.
期刊介绍:
The aim of this journal is to foster the growth of scientific research among Iranian scientists and to provide a medium which brings the fruits of their research to the attention of the world’s scientific community. The journal publishes original research findings – which may be theoretical, experimental or both - reviews, techniques, and comments spanning all subjects in the field of basic sciences, including Physics, Chemistry, Mathematics, Statistics, Biology and Earth Sciences