Recombinant Photorhabdus luminescens subsp. laumondii Quaternary ammonium compound-resistance protein sugE (sugE)
Description
Introduction to sugE Protein
The sugE protein is a member of the small multidrug resistance (SMR) family, initially identified in Escherichia coli as a suppressor of groEL mutations. It has been shown to confer resistance to a subset of toxic quaternary ammonium compounds when overexpressed .
Function and Mechanism
The sugE protein functions as a drug efflux pump, specifically exporting cationic compounds out of bacterial cells. This mechanism is crucial for bacterial survival in environments where quaternary ammonium compounds are present, such as in antiseptic solutions .
Data and Tables
Given the lack of specific data on the recombinant Photorhabdus luminescens subsp. laumondii sugE protein, we can refer to general characteristics of sugE from E. coli:
| Characteristics | Description |
|---|---|
| Gene Name | sugE |
| Function | Drug efflux pump |
| Resistance | Quaternary ammonium compounds |
| Expression Host | E. coli |
| Specific Compounds | Cetylpyridinium, cetyldimethylethyl ammonium, cetrimide |
Future Research Directions
Future studies could focus on cloning and expressing the sugE gene in Photorhabdus luminescens subsp. laumondii to investigate its potential role in conferring resistance to quaternary ammonium compounds in this bacterium. This could involve assessing the protein's structure, function, and efficacy in various environments.
References
-
Overexpression of the Escherichia coli sugE gene confers resistance to a subset of toxic quaternary ammonium compounds.
-
Photorhabdus luminescens and its resistance mechanisms. While not directly related to sugE, Photorhabdus luminescens exhibits resistance mechanisms against antimicrobial peptides, which could provide insights into potential resistance strategies .
-
Recombinant protein production. Techniques for producing recombinant proteins in E. coli could be applied to express sugE in Photorhabdus luminescens subsp. laumondii if the gene is cloned and introduced into this host .
Product Specs
Note: While we prioritize shipping the format currently in stock, please specify your format preference in order notes for customized fulfillment.
Note: Standard shipping includes blue ice packs. Dry ice shipping requires prior arrangement and incurs additional charges.
The tag type is determined during production. If a specific tag type is required, please inform us, and we will prioritize its development.
Target Background
Function: Guanidinium ion exporter. This protein couples guanidinium export to the proton motive force, exchanging one guanidinium ion for two protons.
KEGG: plu:plu4129
Q&A
What is the biological role of the sugE protein in Photorhabdus luminescens subsp. laumondii?
The sugE protein belongs to the small multidrug resistance (SMR) family and is implicated in conferring resistance to toxic quaternary ammonium compounds (QACs). These compounds are commonly used as disinfectants and antiseptics due to their antimicrobial properties. In Photorhabdus luminescens subsp. laumondii, sugE functions as a membrane-associated efflux pump that actively exports QACs from the bacterial cell, thereby reducing intracellular toxicity . This mechanism is essential for bacterial survival in environments where QACs are prevalent, such as soil ecosystems inhabited by nematodes.
The biological significance of sugE extends beyond mere resistance; it plays a role in the bacterium's symbiotic relationship with nematodes (Heterorhabditis species) and its pathogenicity against insect hosts. By protecting bacterial cells from chemical stressors, sugE ensures the survival and replication of Photorhabdus luminescens, which serves as a food source for nematodes during their reproductive cycles . This intricate interplay highlights the evolutionary adaptation of sugE in facilitating both symbiosis and pathogenicity.
How can recombinant expression systems be optimized for producing sugE protein?
Recombinant expression of proteins like sugE often encounters challenges such as low yield, improper folding, or aggregation. To optimize production:
Host Selection
The choice of host organism significantly impacts protein yield and functionality. Commonly used hosts include Escherichia coli, yeast (Saccharomyces cerevisiae), and insect cells (Spodoptera frugiperda). Among these, E. coli remains the most popular due to its rapid growth and well-characterized genetic systems .
Codon Optimization
Codon usage bias can hinder efficient translation of recombinant genes. Tools like TIsigner employ synonymous codon substitutions to enhance mRNA accessibility at translation initiation sites, thereby improving expression outcomes . For example, modifying up to nine codons near the initiation site has been shown to increase protein production while minimizing detrimental effects on host cell growth.
Fusion Tags
Fusion tags such as His-tags or GST-tags facilitate purification and improve solubility but may require removal post-purification to ensure native functionality.
Induction Conditions
Induction parameters such as temperature, inducer concentration (e.g., IPTG), and duration must be optimized. Lower temperatures often favor proper folding of membrane proteins like sugE by reducing aggregation risks .
Membrane Environment
SugE's function as an efflux pump necessitates its integration into lipid bilayers. Co-expression with chaperones or lipoproteins may enhance membrane insertion efficiency.
What experimental approaches can be used to characterize the functional activity of sugE?
Characterizing sugE's activity involves assessing its ability to confer resistance against QACs through efflux mechanisms:
Drug Resistance Assays
Growth inhibition assays can quantify resistance levels by comparing bacterial growth rates in media containing varying concentrations of QACs. Overexpression of sugE typically results in higher survival rates under toxic conditions .
Efflux Pump Assays
Fluorescent dyes such as ethidium bromide or Nile Red serve as substrates for efflux pumps. Monitoring dye accumulation inside cells using fluorescence spectroscopy provides insights into pump activity.
Protein Localization Studies
Immunofluorescence microscopy or GFP-tagging techniques can visualize sugE's localization within membranes. These methods confirm its integration into lipid bilayers—a prerequisite for efflux function .
Structural Analysis
X-ray crystallography or cryo-electron microscopy elucidates the three-dimensional structure of sugE, revealing key domains involved in substrate binding and transport.
How does environmental stress influence the expression and activity of sugE?
Environmental factors such as temperature, pH, and chemical exposure significantly impact sugE expression:
Temperature Effects
Studies have shown that growth temperature influences bacterial infectivity and survival mechanisms . For instance, Photorhabdus luminescens strains grown at lower temperatures exhibit enhanced resistance profiles due to increased expression of stress-responsive genes like sugE.
Chemical Stressors
Exposure to QACs triggers upregulation of efflux pumps, including sugE. This adaptive response ensures bacterial survival under chemical stress.
Symbiotic Context
In symbiotic environments with nematodes, sugE expression may be modulated by host-derived signals or nutrient availability.
What are the implications of sugE-mediated resistance for antibiotic stewardship?
SugE's ability to export QACs raises concerns about cross-resistance to other antimicrobial agents:
Multidrug Resistance
Efflux pumps like sugE often exhibit broad substrate specificity, potentially contributing to multidrug resistance phenotypes. This underscores the importance of monitoring efflux-mediated resistance mechanisms in clinical isolates .
Impact on Disinfectant Efficacy
Overuse of QAC-based disinfectants may select for resistant strains harboring efflux pumps like sugE, diminishing their effectiveness over time.
How do genetic variations in sugE affect its functionality?
Genetic polymorphisms within the sugE gene can alter its substrate specificity or efflux efficiency:
Mutational Analysis
Site-directed mutagenesis allows researchers to pinpoint amino acid residues critical for substrate binding or transport activity.
Comparative Genomics
Sequence comparisons across different strains of Photorhabdus luminescens reveal conserved motifs essential for efflux function.
Can computational modeling predict the behavior of recombinant sugE?
Computational tools such as molecular dynamics simulations provide valuable insights into protein behavior:
Structural Predictions
Homology modeling based on known SMR family structures predicts binding sites and conformational changes during substrate transport.
Expression Outcomes
Machine learning algorithms like TIsigner simulate codon optimization scenarios to forecast recombinant expression success rates .
What challenges arise in studying membrane proteins like sugE?
Membrane proteins pose unique challenges due to their hydrophobic nature:
Solubility Issues
SugE's integration into lipid bilayers complicates purification efforts. Detergents or liposomes are often required for stabilization.
Functional Validation
Reconstituting functional efflux pumps in artificial membranes demands precise lipid composition matching native environments.
How does geographic variation influence bacterial resistance profiles?
Geographic differences among Photorhabdus luminescens strains impact their resistance mechanisms:
Strain-Specific Traits
Clinical isolates from different regions exhibit variable infectivity and survival rates under stress conditions . For example, European strains show reduced association with mammalian cells compared to North American isolates.
