ArticleThe Journal of biological chemistry2011
Preparative scale cell-free production and quality optimization of MraY homologues in different expression modes.
Article in The Journal of biological chemistry, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 22 papers.
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Who cites it
22 citing papers in PubMed, 60 citations in OpenAlex.
- The Shape of Things to Come: α-Helical Membrane Protein Folding on the Ribosome.Chemical reviews · 2026Review
- Cell-free expression system: a promising platform for bacteriophage production and engineering.Microbial cell factories · 2025Review
- Cell-Free Expression of a Plant Membrane Protein BrPT2 From Boesenbergia Rotunda.Molecular biotechnology · 2021Article
- Review
- Development of a Cell-Free Optical Biosensor for Detection of a Broad Range of Mercury Contaminants in Water: A Plasmid DNA-Based Approach.ACS omega · 2019Article
- Aminoribosylated Analogues of Muraymycin Nucleoside Antibiotics.Molecules (Basel, Switzerland) · 2018Article
- Analogues of Muraymycin Nucleoside Antibiotics with Epimeric Uridine-Derived Core Structures.Molecules (Basel, Switzerland) · 2018Article
- Rapid and efficient production of cecropin A antibacterial peptide in Escherichia coli by fusion with a self-aggregating protein.BMC biotechnology · 2018Article
- Insights into the Target Interaction of Naturally Occurring Muraymycin Nucleoside Antibiotics.ChemMedChem · 2018Article
- Bacterial phosphoglycosyl transferases: initiators of glycan biosynthesis at the membrane interface.Glycobiology · 2017Review
- Article
- Fluorescence-based assay for polyprenyl phosphate-GlcNAc-1-phosphate transferase (WecA) and identification of novel antimycobacterial WecA inhibitors.Analytical biochemistry · 2016Article
- The Membrane Steps of Bacterial Cell Wall Synthesis as Antibiotic Targets.Antibiotics (Basel, Switzerland) · 2016Review
- New Insight into the Catalytic Mechanism of Bacterial MraY from Enzyme Kinetics and Docking Studies.The Journal of biological chemistry · 2016Article
- Lipid Requirements for the Enzymatic Activity of MraY Translocases and in Vitro Reconstitution of the Lipid II Synthesis Pathway.The Journal of biological chemistry · 2016Article
- Muraymycin nucleoside-peptide antibiotics: uridine-derived natural products as lead structures for the development of novel antibacterial agents.Beilstein journal of organic chemistry · 2016Review
- Core Steps of Membrane-Bound Peptidoglycan Biosynthesis: Recent Advances, Insight and Opportunities.Antibiotics (Basel, Switzerland) · 2015Review
- Chemoenzymatic syntheses of water-soluble lipid I fluorescent probes.Tetrahedron letters · 2015Article
- Biosynthesis of a water-soluble lipid I analogue and a convenient assay for translocase I.Analytical biochemistry · 2014Article
- Fluorescent probe for high-throughput screening of membrane protein expression.Protein science : a publication of the Protein Society · 2013Article
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Authors and funding
10 authors at 4 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
MraY translocase catalyzes the first committed membrane-bound step of bacterial peptidoglycan synthesis leading to the formation of lipid I. The essential membrane protein therefore has a high potential as target for drug screening approaches to develop antibiotics against gram-positive as well as gram-negative bacteria. However, the production of large integral membrane proteins in conventional cellular expression systems is still very challenging. Cell-free expression technologies have been optimized in recent times for the production of membrane proteins in the presence of detergents (D-CF), lipids (L-CF), or as precipitates (P-CF). We report the development of preparative scale production protocols for the MraY homologues of Escherichia coli and Bacillus subtilis in all three cell-free expression modes followed by their subsequent quality evaluation. Although both proteins can be cell-free produced at comparable high levels, their requirements for optimal expression conditions differ markedly. B. subtilus MraY was stably folded in all three expression modes and showed highest translocase activities after P-CF production followed by defined treatment with detergents. In contrast, the E. coli MraY appears to be unstable after post- or cotranslational solubilization in detergent micelles. Expression kinetics and reducing conditions were identified as optimization parameters for the quality improvement of E. coli MraY. Most remarkably, in contrast to B. subtilis MraY the E. coli MraY has to be stabilized by lipids and only the production in the L-CF mode in the presence of preformed liposomes resulted in stable and translocase active protein samples.
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