Plasmids are one of the most important genetic tools for basic research and biotechnology, as they enable rapid genetic manipulation. Here we present a novel pBBR1-based plasmid for Methylorubrum extorquens, a model methylotroph that is used for the development of C1-based microbial cell factories. To develop a vector with compatibility to the so far mainly used pCM plasmid system, we transferred the pBBR1-based plasmid pMiS1, which showed an extremely low transformation rate and caused a strong growth defect. Isolation of a suppressor mutant with improved growth led to the isolation of the variant pMis1_1B. Its higher transformation rate and less pronounced growth defect phenotype could be shown to be the result of a mutation in the promotor region of the rep gene. Moreover, cotransformation of pMis1_1B and pCM160 was possible, but the resulting transformants showed stronger growth defects in comparison with a single pMis1_1B transformant. Surprisingly, cotransformants carrying pCM160 and a pMis1_1B derivative containing a mCherry reporter construct showed higher fluorescence levels than strains containing only the pMis1_1B-based reporter plasmids or a corresponding pCM160 derivative. Relative plasmid copy number determination experiments confirmed our hypothesis of an increased copy number of pMis1_1B in the strain carrying both plasmids. Despite the slight metabolic burden caused by pMis1_1B, the plasmid strongly expands the genetic toolbox for M. extorquens.
Download full-text PDF |
Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9531332 | PMC |
| http://dx.doi.org/10.1002/mbo3.1325 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
A novel engineered strain of Methylorubrum extorquens for methylotrophic production of glycolic acid.
Microb Cell Fact
December 2024
Fraunhofer Institute for Interfacial Engineering and Biotechnology, Straubing Branch BioCat, Schulgasse 11a, Straubing, Germany.
The conversion of CO into methanol depicts one of the most promising emerging renewable routes for the chemical and biotech industry. Under this regard, native methylotrophs have a large potential for converting methanol into value-added products but require targeted engineering approaches to enhance their performances and to widen their product spectrum. Here we use a systems-based approach to analyze and engineer M.
View Article and Find Full Text PDFModulating metal-centered dimerization of a lanthanide chaperone protein for separation of light lanthanides.
Proc Natl Acad Sci U S A
November 2024
Department of Chemistry, The Pennsylvania State University, University Park, PA 16802.
Elucidating details of biology's selective uptake and trafficking of rare earth elements, particularly the lanthanides, has the potential to inspire sustainable biomolecular separations of these essential metals for myriad modern technologies. Here, we biochemically and structurally characterize () LanD, a periplasmic protein from a bacterial gene cluster for lanthanide uptake. This protein provides only four ligands at its surface-exposed lanthanide-binding site, allowing for metal-centered protein dimerization that favors the largest lanthanide, La.
View Article and Find Full Text PDFEvolutionary engineering of methylotrophic E. coli enables fast growth on methanol.
Nat Commun
October 2024
Institute of Biological Chemistry, Academia Sinica, Taipei City, Taiwan, ROC.
Article Synopsis
- Methanol can be produced from CO or methane, making it a key player in addressing climate change through a methanol economy.
- Researchers developed a new strain of E. coli that grows rapidly, with a doubling time of 3.5 hours, which is competitive with other methylotrophs.
- The study highlights the use of dynamic copy number variation (CNV) in creating synthetic microorganisms and points to their potential for sustainable biotechnological advancements.
Combined application of resveratrol and a ryegrass endophyte in PAH-contaminated soil remediation and its impact on soil microbial communities.
RSC Adv
October 2024
Zhejiang Provincial Key Laboratory of Solid Waste Treatment and Recycling, Zhejiang Engineering Research Center of Non-ferrous Metal Waste Recycling, School of Environmental Science and Engineering, Zhejiang Gongshang University Hangzhou 310012 China
Article Synopsis
- Certain plant endophytes have shown potential in breaking down organic pollutants, specifically polycyclic aromatic hydrocarbons (PAHs), but their effectiveness after culture and in real-world soil remediation is still uncertain.
- A study found that resveratrol enhances the PAH degradation ability of an endophyte called C1 while having little effect on native soil bacteria, suggesting a selective boosting effect.
- The combination of resveratrol and the endophyte C1 in contaminated soil resulted in significantly improved PAH removal rates, indicating a promising new bioremediation method that utilizes the interaction between plant-produced metabolites and endophytic bacteria.
Structure-driven development of a biomimetic rare earth artificial metalloprotein.
Proc Natl Acad Sci U S A
August 2024
Center for Biophysics and Quantitative Biology, University of Illinois, Urbana-Champaign, Urbana, IL 61801.
The 2011 discovery of the first rare earth-dependent enzyme in methylotrophic AM1 prompted intensive research toward understanding the unique chemistry at play in these systems. This enzyme, an alcohol dehydrogenase (ADH), features a La ion closely associated with redox-active coenzyme pyrroloquinoline quinone (PQQ) and is structurally homologous to the Ca-dependent ADH from the same organism. AM1 also produces a periplasmic PQQ-binding protein, PqqT, which we have now structurally characterized to 1.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!