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

IF 1.4 4区 综合性期刊 Q2 MULTIDISCIPLINARY SCIENCES
Nasira Rafi, Rabia Abbas, Nadeem Ahmed, Saad Tahir, Muhammad Akram, Mohsin Ahmad Khan, Nao Akusa Fujimura, Mariam Shafique, Kausar Malik
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引用次数: 0

摘要

治疗用蛋白质需要大量生产,因为它们在医药市场上的份额越来越大。人 beta 干扰素(hIFN-β)具有抗病毒、抗增殖和免疫调节的特性,可用于治疗各种疾病。考虑到其临床应用,需要开发一种简单、经济、易扩展的生产系统,以高产、高质量地生产 rhIFN-β-1b 蛋白。改良生产方法的尝试一直在进行。以前的大多数报道都提出用异丙基-β-D-1-硫代吡喃半乳糖苷(IPTG)在大肠杆菌中表达 rhIFN-β-1b,这种方法成本高且不能代谢。因此,本研究采用廉价的自动诱导培养基来优化 rhIFN-β-1b 的表达。研究使用了玉米浸出液(CSL),并优化了不同浓度的玉米浸出液。此外,还使用不同浓度的甘油对培养基的碳源进行了优化。最后,进行了一次 10 升发酵(10% CSL 和 2.5% 甘油),产生了 65 克湿生物质和 15 克包涵体(湿)。通过一步离子交换色谱法(IEX)对纯化方案进行了优化,使目标蛋白质的纯度达到 99%。通过抑制病毒性水泡性口炎(VSV)对马丁达比牛肾(MDBK)细胞的细胞病理学作用,评估了所需蛋白质的生物活性。此外,还研究了该蛋白对不同乳腺癌细胞系(HCC- 1954、MCF- 7 和 MDA- MB- 231)的细胞毒性。这是首次报道利用 CSL 作为自动诱导介质来表达和纯化 rhIFN-β-1b。因此,建议的方法可用于生产 rhIFN-β-1b 和其他重组蛋白,不仅简单,而且成本效益高。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Corn Steep Liquor as an Auto-Induction Medium for the Production of rhIFN-β-1b Protein in Escherichia coli

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.

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来源期刊
CiteScore
4.00
自引率
5.90%
发文量
122
审稿时长
>12 weeks
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
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