Comparing the Soluble Form of Recombinant Human Insulin-like Growth Factor-1 (rhIGF-1) in Escherichia coli Using Thioredoxin as Fused and Co-expressed Protein.

IF 1 4区生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Protein and Peptide Letters Pub Date : 2024-01-01 DOI:10.2174/0109298665314267240624091046

Sara Hemmati, Parvaneh Maghami, Javad Ranjbari, Maryam Tabarzad

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Abstract

Introduction: Insulin-like growth factor-1 (IGF-1) is a single-chain polypeptide with various physiological functions. Escherichia coli is one of the most desirable hosts for recombinant protein production, especially for human proteins whose post-translation modifications are not essential for their bioactivity, such as hIGF-1.

Objectives: In this study, bacterial thioredoxin (Trx) was studied as a fused and non-fused protein to convert the insoluble form of recombinant human IGF-1 (rhIGF-1) to its soluble form in E. coli.

Methods: The rhIGF-1 was expressed in the E. coli Origami strain in the form of fused-Trx. It was co-expressed with Trx and then purified and quantified. In the next step, the biological activity of rhIGF-1 was evaluated by alkaline phosphatase (ALP) activity assay in human adipose- derived stem cells (hASCs) regarding the differentiation enhancement effect of IGF-1 through the osteogenic process.

Results: Results showed that Trx in both the fused and non-fused forms had a positive effect on the production of the soluble form of rhIGF-1. A significant increase in ALP activity in hASCs after rhIGF-1 treatment was observed, confirming protein bioactivity.

Conclusion: It was strongly suggested that the overproduction of Trx could increase the solubility of co-expressed recombinant proteins by changing the redox state in E. coli cells.

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利用硫氧还蛋白作为融合蛋白和共表达蛋白比较大肠杆菌中重组人胰岛素样生长因子-1（rhIGF-1）的可溶性形式。

简介胰岛素样生长因子-1（IGF-1）是一种单链多肽，具有多种生理功能。大肠杆菌是生产重组蛋白质最理想的宿主之一，尤其是那些翻译后修饰对其生物活性并不重要的人类蛋白质，如 hIGF-1：本研究研究了细菌硫氧还蛋白（Trx）作为融合蛋白和非融合蛋白在大肠杆菌中将重组人 IGF-1 （rhIGF-1）的不溶性形式转化为可溶性形式：方法：在大肠杆菌 Origami 菌株中以融合-Trx 的形式表达 rhIGF-1。方法：在大肠杆菌 Origami 菌株中以融合-Trx 的形式表达 rhIGF-1，并与 Trx 共同表达，然后进行纯化和定量。下一步，通过碱性磷酸酶（ALP）活性测定评估了rhIGF-1在人脂肪来源干细胞（hASCs）中的生物活性，以了解IGF-1通过成骨过程增强分化的作用：结果表明，融合型和非融合型Trx对可溶性rhIGF-1的产生有积极影响。rhIGF-1处理后，hASCs的ALP活性明显增加，证实了蛋白质的生物活性：结论：这强烈表明，Trx 的过度产生可通过改变大肠杆菌细胞中的氧化还原状态来增加共表达重组蛋白的溶解度。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Protein and Peptide Letters 生物-生化与分子生物学

CiteScore

2.90

自引率

0.00%

发文量

审稿时长

2 months

期刊介绍： Protein & Peptide Letters publishes letters, original research papers, mini-reviews and guest edited issues in all important aspects of protein and peptide research, including structural studies, advances in recombinant expression, function, synthesis, enzymology, immunology, molecular modeling, and drug design. Manuscripts must have a significant element of novelty, timeliness and urgency that merit rapid publication. Reports of crystallization and preliminary structure determination of biologically important proteins are considered only if they include significant new approaches or deal with proteins of immediate importance, and preliminary structure determinations of biologically important proteins. Purely theoretical/review papers should provide new insight into the principles of protein/peptide structure and function. Manuscripts describing computational work should include some experimental data to provide confirmation of the results of calculations. Protein & Peptide Letters focuses on: Structure Studies Advances in Recombinant Expression Drug Design Chemical Synthesis Function Pharmacology Enzymology Conformational Analysis Immunology Biotechnology Protein Engineering Protein Folding Sequencing Molecular Recognition Purification and Analysis