Recombinant Shewanella woodyi ATP synthase subunit c (atpE)
Description
Functional Role in ATP Synthase
Subunit c forms a cylindrical oligomer (c-ring) in the F₀ sector, facilitating proton translocation across the membrane. This rotation drives ATP synthesis in the F₁ sector through mechanical coupling . Key functional insights include:
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Proton Translocation:
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Structural Specificity:
Research Applications and Suppliers
Recombinant Shewanella woodyi atpE is utilized in enzymatic studies, drug discovery, and structural biology.
Applications
Suppliers
Research Findings and Implications
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Non-Redundant Isoforms:
Subunit c isoforms (e.g., P1, P2 in human mitochondria) exhibit distinct roles in respiratory chain assembly and function. Silencing isoforms individually impaired ATP synthesis and cytochrome oxidase activity, underscoring functional specificity . -
Drug Resistance and Targets:
Mutations in subunit c (e.g., Ala63→Met in Mycobacterium) confer resistance to TMC207, a drug targeting the c-ring . This highlights the subunit’s potential as a therapeutic target in tuberculosis and other bacterial infections. -
Recombinant Production Advances:
Codon optimization and MBP fusion strategies enable soluble expression of hydrophobic c-subunits in E. coli, overcoming traditional challenges in membrane protein production .
Product Specs
Please note that we will prioritize shipping the format currently available in stock. However, if you have a specific format preference, kindly include your request in the order remarks, and we will accommodate your needs to the best of our ability.
Standard shipping for our proteins includes normal blue ice packs. If you require dry ice shipping, please inform us in advance. Additional charges may apply.
Generally, the shelf life for liquid forms is 6 months at -20°C/-80°C. For lyophilized forms, the shelf life is 12 months at -20°C/-80°C.
Target Background
KEGG: swd:Swoo_4903
STRING: 392500.Swoo_4903
Q&A
What is Shewanella woodyi ATP synthase subunit c (atpE)?
Shewanella woodyi ATP synthase subunit c (atpE) is a critical component of the ATP synthase complex in S. woodyi bacteria. This protein functions as part of the membrane-embedded Fo domain of ATP synthase, forming the c-ring structure that is essential for the rotary mechanism driving ATP synthesis. While specific information on S. woodyi atpE is limited, it likely shares functional similarities with other bacterial ATP synthase c subunits, participating in ion translocation across the membrane to generate the rotational force necessary for ATP production .
What are the structural characteristics of ATP synthase c subunits in bacteria like Shewanella?
ATP synthase c subunits in bacteria typically display one of two structural arrangements:
| Type | Structure | Examples | Ion Binding Sites |
|---|---|---|---|
| F-type (common) | One hairpin structure | E. coli, P. modestum | One ion binding site per hairpin |
| V-type/hybrid | V-type c subunit | Some anaerobic archaea, A. woodii (hybrid) | May have modified ion binding characteristics |
The c subunit in S. woodyi likely adopts the F-type structure common to most bacteria, though specific structural studies would be required for confirmation .
What expression systems are most effective for producing recombinant S. woodyi ATP synthase subunit c?
For successful expression of recombinant S. woodyi ATP synthase subunit c, heterologous expression in E. coli is typically the method of choice, as demonstrated with other ATP synthases. Based on studies of similar proteins, recommended approaches include:
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Using pET expression vectors with T7 promoter systems
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Expression in E. coli strains designed for membrane protein production (e.g., C43(DE3))
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Addition of a purification tag (His-tag) for subsequent affinity chromatography
Expression should be optimized with varying induction temperatures (typically 18-30°C) and inducer concentrations to maximize yield while ensuring proper folding of this membrane protein .
How can researchers measure ATP synthase activity in reconstituted systems?
ATP synthase activity can be measured using the following methodologies:
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Proteoliposome Reconstitution: After purification, ATP synthase should be reconstituted into liposomes to create proteoliposomes that mimic the native membrane environment.
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Artificial Potential Generation: Create an electrochemical gradient by:
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Establishing a K+ diffusion potential using valinomycin (for Δψ)
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Creating an ion concentration gradient (for ΔpNa or ΔpH)
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ATP Synthesis Measurement:
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Add ADP and Pi to the proteoliposomes
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Measure ATP production using luciferase-based assays or coupled enzyme assays
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Monitor synthesis rates over time (typically linear for approximately 2 minutes)
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This approach allows testing of ATP synthesis under varying conditions of membrane potential and ion gradients .
How do energetic thresholds for ATP synthesis compare between different bacterial ATP synthases?
Energetic thresholds for ATP synthesis vary significantly between different bacterial ATP synthases, which has important implications for understanding bacterial adaptations to different environments:
| Organism | ATP Synthase Type | Energetic Threshold | Can Use Δψ Alone | Can Use ΔpH/ΔpNa Alone |
|---|---|---|---|---|
| E. callanderi | A₁AO with V-type c | 87 mV | Yes | Yes |
| A. woodii | F₁FO (hybrid rotor) | 90 mV | Yes | No |
| P. modestum | Na⁺-F₁FO | 120 mV | No | No |
| E. coli | H⁺-F₁FO | 150 mV | No | No |
S. woodyi ATP synthase has not been specifically characterized in terms of these parameters, but as a marine bacterium, it might have adaptations for functioning in its natural environment. Researchers should consider these comparative data when designing experiments to characterize S. woodyi ATP synthase .
How might S. woodyi ATP synthase be involved in biofilm formation and environmental adaptation?
While not directly studied for S. woodyi, research on related Shewanella species provides important insights into potential roles of ATP synthase in biofilm formation:
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Energy Production During Biofilm Development: ATP synthase likely provides the energy required for the transition from planktonic to biofilm lifestyles.
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Environmental Sensing: In S. oneidensis, the Arc two-component system (including ArcA) regulates responses to changes in oxygen levels. This system may interact with energy production mechanisms, including ATP synthase, to modulate biofilm formation based on environmental conditions .
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Metabolic Adaptation: ATP synthase activity may be regulated differently during biofilm formation to accommodate the altered metabolic needs of biofilm cells compared to planktonic cells.
Future research specifically examining S. woodyi ATP synthase in the context of biofilm formation would be valuable for understanding its environmental adaptations .
What can phylogenetic analysis tell us about the evolution of ATP synthase subunit c in Shewanella species?
Phylogenetic analysis of ATP synthase subunit c provides valuable insights into the evolutionary history and adaptations of Shewanella species:
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Evolutionary Conservation: The c subunit is generally well-conserved due to its critical role in energy production, though variations exist in ion specificity and structural organization.
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Lateral Gene Transfer: Some unusual ATP synthase configurations may result from lateral gene transfer events, as suggested by the presence of V-type c subunits in some bacteria that normally contain F-type ATP synthases.
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Environmental Adaptation: Variations in the c subunit sequence may reflect adaptations to different environmental conditions, including marine environments in the case of S. woodyi.
Comparative analysis between Shewanella species and other bacteria can reveal how ATP synthase has evolved to function in various ecological niches .
How do the structural characteristics of ATP synthase c subunit influence its function in different organisms?
The structural characteristics of the ATP synthase c subunit have profound effects on function:
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Ion Binding Sites: The number and nature of ion binding sites determine the ion:ATP ratio and therefore the thermodynamic efficiency of ATP synthesis.
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Structural Organization:
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F-type c subunits typically contain two transmembrane helices forming one hairpin with one ion binding site
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V-type c subunits have four transmembrane helices with potentially different ion binding properties
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Oligomeric Ring Structure: The number of c subunits in the ring varies between species (9-15 subunits) and affects the bioenergetics of ATP synthesis.
For S. woodyi, the specific structural features of its ATP synthase c subunit would determine its functional properties, though detailed structural studies are needed to characterize these features fully .
What are the key considerations for designing experiments to characterize S. woodyi ATP synthase function?
When designing experiments to characterize S. woodyi ATP synthase function, researchers should consider:
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Expression and Purification Strategy:
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Select appropriate expression systems for membrane proteins
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Develop effective purification protocols that maintain protein stability
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Consider using affinity tags that can be removed for functional studies
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Functional Reconstitution:
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Reconstitute purified ATP synthase into liposomes with appropriate lipid composition
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Establish methods to generate defined ion gradients and membrane potentials
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Develop reliable assays for measuring ATP synthesis and hydrolysis
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Comparative Analysis:
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Include well-characterized ATP synthases (e.g., from E. coli) as benchmarks
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Test function under a range of conditions to identify optimal operating parameters
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Consider physiologically relevant conditions based on S. woodyi's natural habitat
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Structural Studies:
How can researchers investigate the relationship between S. woodyi ATP synthase and electron transport systems?
Investigating the relationship between S. woodyi ATP synthase and electron transport systems requires integrated approaches:
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Genetic Approaches:
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Create knockout mutants of ATP synthase components and electron transport components
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Use whole-genome knockout collections similar to those developed for S. oneidensis
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Employ complementation studies to verify gene functions
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Bioenergetic Measurements:
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Measure membrane potential and ion gradients in vivo using fluorescent probes
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Determine ATP synthesis rates under different electron donor and acceptor conditions
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Assess the effects of inhibitors specific to different components of the electron transport chain
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In vitro Reconstitution:
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Co-reconstitute purified ATP synthase with components of the electron transport chain
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Measure electron transfer and ATP synthesis activities in the reconstituted system
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This integrated approach would provide insights into how S. woodyi coordinates electron transport and ATP synthesis, particularly in the context of its unique respiratory capabilities and environmental adaptations .
