High-level production of heme-containing holoproteins in Escherichia coli

Applied Microbiology and Biotechnology
Y JungY Lee

Abstract

The expression of recombinant protein is essential for the investigation of the functions and properties of heme-containing protein as an electron carrier. For the expression of fully active recombinant protein, conversion of the expressed apoprotein into holoprotein is the most important and difficult problem. In this study, a system was developed for the production of heme-containing protein in a pure, recombinant holoprotein form, using the bovine cytochrome b5 tryptic fragment and Escherichia coli bacterioferritin as heterologous and homologous heme-containing model proteins, respectively. This system is based on the slow synthesis of recombinant apoprotein, which can maintain the balanced consumption of amino acids between protein synthesis and heme synthesis, so that the synthesized apoprotein continues to act as a heme sink. From a 1-1 culture, 15 mg of cytochrome b5 and 40 mg of bacterioferritin were purified as pure holoprotein forms. Our expression system provides a rapid and simple method for obtaining large quantities of the active holo-form of heme-containing proteins.

Citations

Sep 11, 2007·Applied Microbiology and Biotechnology·Irene NeophytouDavid Barlow
Nov 4, 2010·The Journal of Microbiology·Marta A Almirón, Rodolfo A Ugalde
Mar 25, 2004·Protein Expression and Purification·Cornelius L Varnado, Douglas C Goodwin
Jul 4, 2006·Trends in Biotechnology·Barry J RyanCiarán O'Fágáin
Jan 2, 2014·Protein Expression and Purification·Karen HänelAngelika Loidl-Stahlhofen
Apr 25, 2017·Biotechnology and Bioengineering·Yongkun LvJian Chen
Jun 24, 2017·Preparative Biochemistry & Biotechnology·Changqing LiuHuizhou Liu
Jul 14, 2010·Journal of Molecular Modeling·Ying-Wu LinLi-Fu Liao
Jul 31, 2021·Fish Physiology and Biochemistry·Muhammad Mehedi HasanYoshihiro Ochiai
Oct 15, 2014·ACS Medicinal Chemistry Letters·Shingo TojoYoshiaki Isobe

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