Saronom Silaban, Murniaty Simorangkir, Iman Permana Maksum, Khomaini Hasan

Pengaruh Suhu pada Ekspresi Pretrombin-2 Manusia Rekombinan di Escherichia coli ER2566

Introduction

Pengaruh suhu pada ekspresi pretrombin-2 manusia rekombinan di escherichia coli er2566. Temukan pengaruh suhu induksi terhadap ekspresi pretrombin-2 manusia rekombinan (rhPT-2) dalam E. coli ER2566. Pelajari bagaimana 22°C menjadi suhu optimum untuk fraksi larut.

Abstract

Human recombinant prethrombin-2 (rhPT-2) was expressed in Escherichia coli. Previous studies reported that the expression of rhPT-2 in E. coli tends to formed inclusion body. This study aims to determine the effect of temperature on the expression of rhPT-2 in E. coli ER2566. This study begins with the isolation of recombinant plasmids, competent cell preparation and transformation of competent cells of E. coli ER2566 strain, and rhPT-2 gene expression with various induction temperatures 12, 18, 22 and 30°C. Characterization results showed that the induction temperature of 22°C was the optimum temperature of rhPT-2 gene expression as a soluble fraction in E. coli ER2566 with IPTG concentration 0.1 mM. These results prove that the temperature affects the expression of rhPT-2 in E. coli ER2566. [EFFECT OF TEMPERATURE ON RECOMBINANT HUMAN PRETROMBIN-2 EXPRESSION IN ESCHERICHIA COLI ER2566] (J. Sains Indon., 42(2): 25-30, 2018)Keywords:Induction Temperature, Soluble Fraction, Prethrombin-2, E. coli ER2566

Review

This study capably addresses a common and significant challenge in recombinant protein expression: the formation of insoluble inclusion bodies. By focusing on human recombinant prethrombin-2 (rhPT-2) expression in *Escherichia coli* ER2566, the authors aimed to systematically investigate the effect of induction temperature on protein solubility. This is a pertinent area of research given the therapeutic potential of many recombinant proteins, where soluble and functional yield is paramount. The initial premise, acknowledging prior difficulties with rhPT-2 solubility in *E. coli*, sets a clear and practical goal for the investigation. The experimental approach described involves standard molecular biology techniques, including plasmid isolation, competent cell preparation, and transformation, followed by expression trials at various induction temperatures (12, 18, 22, and 30°C). A key strength of the study lies in its focused empirical determination of an optimal parameter. The finding that 22°C induction temperature, coupled with 0.1 mM IPTG, significantly enhanced the expression of rhPT-2 in its soluble form is a valuable and actionable result. This specific optimization provides a practical starting point for researchers aiming to produce soluble rhPT-2 in this *E. coli* strain. While the abstract clearly identifies an optimal temperature for soluble rhPT-2 expression, it leaves some critical questions unanswered that would ideally be addressed in the full manuscript. For instance, the abstract states "characterization results showed," but only mentions solubility; there is no indication of the total protein yield achieved, nor any assessment of the expressed protein's biological activity or proper folding, which are crucial for a functional protein like prethrombin-2. Furthermore, while temperature and IPTG concentration are mentioned, the abstract does not elaborate on the optimization process for IPTG or induction duration, which are also vital for maximizing soluble yield. A quantitative comparison showing the *extent* of improvement in soluble fraction at 22°C compared to other temperatures, especially higher temperatures that typically lead to inclusion bodies, would further strengthen the claims. Despite these areas for potential elaboration, the study provides a useful contribution to optimizing bioprocesses for recombinant protein production.

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