Engineered Chimera Protein Constructs to Facilitate the Production of Heterologous Transmembrane Proteins in E. coli

• Published in: International journal of molecular sciences Vol. 25; no. 4; p. 2354
• Main Authors: Ogunbowale, Adeyemi, Georgieva, Elka R
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 16-02-2024
• Subjects: Amino acids; Cell Membrane - metabolism; E coli; E. coli expression host of heterologous transmembrane proteins; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - metabolism; Ligands; Localization; Maltose-Binding Proteins - genetics; Maltose-Binding Proteins - metabolism; Membrane Proteins - metabolism; Membranes; Peptides; Physiology; Plasma; protein engineering; Protein expression; Protein Sorting Signals; Proteins; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; soluble transmembrane proteins; transmembrane protein fusion strategies; Yeast; protein engineering; E. coli expression host of heterologous transmembrane proteins; transmembrane protein fusion strategies; soluble transmembrane proteins
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms25042354

To delve into the structure-function relationship of transmembrane proteins (TMPs), robust protocols are needed to produce them in a pure, stable, and functional state. Among all hosts that express heterologous TMPs,
has the lowest cost and fastest turnover. However, many of the TMPs expressed in
are misfolded. Several strategies have been developed to either direct the foreign TMPs to
's membrane or retain them in a cytosolic soluble form to overcome this deficiency. Here, we summarize protein engineering methods to produce chimera constructs of the desired TMPs fused to either a signal peptide or precursor maltose binding protein (pMBP) to direct the entire construct to the periplasm, therefore depositing the fused TMP in the plasma membrane. We further describe strategies to produce TMPs in soluble form by utilizing N-terminally fused MBP without a signal peptide. Depending on its N- or C-terminus location, a fusion to apolipoprotein AI can either direct the TMP to the membrane or shield the hydrophobic regions of the TMP, maintaining the soluble form. Strategies to produce G-protein-coupled receptors, TMPs of
, HIV-1 Vpu, and other TMPs are discussed. This knowledge could increase the scope of TMPs' expression in
.