Recombinant Dinoroseobacter shibae ATP synthase subunit b' (atpG)

What is the role of ATP synthase subunit b' in Dinoroseobacter shibae energy metabolism?

ATP synthase subunit b' is a crucial component of the F₀ portion of the F₁F₀-ATP synthase complex in D. shibae. This complex harnesses the proton motive force generated during respiratory and photosynthetic electron transport to synthesize ATP. In D. shibae, ATP synthase functions within a sophisticated metabolic network that includes aerobic respiration, denitrification, and aerobic anoxygenic photosynthesis.

D. shibae utilizes a unique cascade-type regulatory network involving FnrL and three Dnr regulators (DnrD, DnrE, and DnrF) to control transitions between aerobic and anaerobic metabolism . The ATP synthase complex is likely regulated as part of this network, as it must function efficiently under changing oxygen conditions. Under aerobic conditions, the proton gradient for ATP synthesis is primarily generated through aerobic respiration, while under anaerobic conditions, denitrification pathways become the main source of the proton motive force.

How does the structure of D. shibae ATP synthase b' subunit compare to other bacterial homologs?

The ATP synthase b' subunit in D. shibae shares structural similarities with homologs from other alphaproteobacteria but also possesses unique features reflecting its adaptation to marine environments and metabolic versatility. While the core domain architecture is conserved, D. shibae's b' subunit contains specific amino acid variations that may contribute to the enzyme's stability and function under the organism's versatile energy-generating conditions.

The protein likely contains a membrane-spanning N-terminal domain and a cytoplasmic C-terminal domain that interacts with the F₁ portion of the ATP synthase complex. These structural features enable the b' subunit to function as part of the peripheral stalk, connecting the membrane-embedded F₀ portion with the catalytic F₁ portion of the ATP synthase.

What expression systems have been successfully used for recombinant production of D. shibae atpG?

For recombinant expression of D. shibae ATP synthase subunit b', E. coli-based expression systems have proven most effective. Successful expression typically involves:

• Codon optimization of the atpG gene for E. coli expression

• Fusion with affinity tags (e.g., His₆, Strep-tag II) for purification

• Use of expression vectors with tunable promoters (T7, tac)

• Expression in specialized E. coli strains (BL21(DE3), C41(DE3), C43(DE3)) designed for membrane protein expression

• Optimization of induction conditions (temperature, IPTG concentration, induction time)

The membrane-associated nature of the b' subunit presents challenges for recombinant expression. Using E. coli C41(DE3) or C43(DE3) strains at lower induction temperatures (16-18°C) often improves yield and solubility of the recombinant protein.

How can researchers verify the functionality of recombinant D. shibae ATP synthase subunit b'?

Verification of recombinant D. shibae ATP synthase subunit b' functionality requires multiple approaches:

• Structural integrity assessment:

  • Circular dichroism spectroscopy to confirm secondary structure

  • Size-exclusion chromatography to evaluate oligomeric state

  • Limited proteolysis to assess proper folding

• Binding studies:

  • Pull-down assays with other ATP synthase subunits

  • Surface plasmon resonance to measure binding kinetics

  • Isothermal titration calorimetry for thermodynamic parameters

• Functional complementation:

  • Transformation of b' subunit-deficient bacterial strains

  • Measurement of ATP synthesis rates in complemented strains

  • Assessment of growth under different metabolic conditions

• Reconstitution experiments:

  • Incorporation into liposomes with other ATP synthase subunits

  • Proton pumping assays using pH-sensitive fluorescent dyes

  • ATP synthesis assays in reconstituted systems

These methodological approaches provide comprehensive evaluation of the recombinant protein's structural integrity and functional capacity.

How does D. shibae ATP synthase activity adapt during the transition between aerobic and anaerobic metabolism?

D. shibae demonstrates remarkable metabolic flexibility, transitioning between aerobic respiration, anaerobic denitrification, and aerobic anoxygenic photosynthesis. ATP synthase activity during these transitions involves sophisticated regulatory mechanisms:

During the aerobic-to-anaerobic transition, D. shibae employs a cascade-type regulatory network involving FnrL (an oxygen-sensing regulator with an Fe-S cluster) and three Dnr regulators (DnrD, DnrE, and DnrF) . This regulatory network controls electron transport chain components and potentially modulates ATP synthase activity as follows:

• Oxygen sensing and initial response:

  • FnrL acts as the primary oxygen sensor through its oxygen-sensitive Fe-S cluster

  • Upon oxygen limitation, active FnrL induces expression of high-affinity cbb₃-type cytochrome c oxidase and represses low-affinity aa₃-type cytochrome

  • These changes in electron transport chain composition affect proton motive force generation

• Regulatory cascade progression:

  • FnrL activates expression of dnrE and dnrF regulator genes

  • DnrD, likely sensing NO via a heme cofactor, co-induces genes for denitrification

  • DnrE controls genes for a putative Na⁺/H⁺ antiporter, potentially affecting ion gradients used by ATP synthase

• Coordinated adjustment of energy metabolism:

  • Formation of electron-donating primary dehydrogenases is coordinated by FnrL and DnrE

  • ATP synthase activity must adapt to changes in proton motive force generation

This regulatory network ensures optimal ATP synthase function across different metabolic modes, maintaining energy production efficiency despite changing environmental conditions.

What role might ATP synthase play in the Jekyll and Hyde interaction between D. shibae and dinoflagellates?

The Jekyll and Hyde interaction between D. shibae and the dinoflagellate Prorocentrum minimum involves a mutualistic phase followed by a pathogenic phase . ATP synthase may play critical roles in both phases:

Mutualistic Phase:

• ATP synthase provides energy for vitamin B₁₂ and B₇ synthesis, which D. shibae supplies to its algal host

• Energy from ATP synthase powers flagellar motility, essential for establishing symbiosis, as demonstrated by the inability of flagella-deficient mutants to stimulate P. minimum growth

• ATP synthase activity may be modulated by the CtrA phosphorelay system, which controls flagella biosynthesis and is required for mutualism

Transition to Pathogenic Phase:

• Quorum sensing systems, potentially influencing ATP synthase regulation, show growth phase-dependent changes in expression during co-culture

• Cell density-dependent pathogenicity suggests metabolic shifts potentially involving ATP synthase

• The 191 kb plasmid of D. shibae, essential for pathogenicity , may encode factors that influence energy metabolism and ATP synthase function

Research approaches to investigate this role:

• Comparative proteomics of ATP synthase subunits during mutualistic versus pathogenic phases

• Metabolic flux analysis tracking ATP production and consumption in co-culture

• Construction of atpG mutants with altered expression levels to assess effects on both mutualistic and pathogenic interactions

How does blue light exposure affect ATP synthase activity in D. shibae?

D. shibae possesses a blue light-dependent LOV-protein (LdaP) that acts as an antirepressor of the PpsR repressor, regulating photosynthetic gene clusters . This photosensory system likely influences ATP synthase activity through several mechanisms:

• Photosynthetic gene regulation:

  • Blue light activates LdaP, which inhibits PpsR repressor activity

  • PpsR normally represses the bchFNBHLM operon involved in bacteriochlorophyll biosynthesis

  • Under blue light, derepression occurs, leading to increased photosynthetic apparatus formation

• Metabolic consequences:

  • Enhanced photosynthetic capacity under blue light generates additional proton motive force

  • ATP synthase utilizes this increased proton gradient for ATP production

  • Transcriptional regulation of ATP synthase genes may occur in coordination with photosynthetic genes

• Experimental evidence:

  • β-galactosidase activity measurements using bchF-lacZ reporter fusions show significantly higher expression under blue light (467 nm) compared to dark conditions in wild-type D. shibae

  • In the ppsR::Tn mutant strain, expression levels are constitutively high regardless of light conditions

  • These findings suggest coordinated regulation of energy-generating systems

Researchers investigating this relationship should consider:

• Measuring ATP synthase activity directly under different light conditions

• Performing transcriptomic and proteomic analyses of ATP synthase components following blue light exposure

• Constructing reporter gene fusions for atpG to monitor expression in response to light

What technical challenges exist in structural studies of recombinant D. shibae ATP synthase subunit b'?

Structural characterization of recombinant D. shibae ATP synthase subunit b' presents several technical challenges:

• Membrane association and hydrophobicity:

  • The N-terminal membrane-spanning domain creates solubility issues

  • Detergent selection is critical for maintaining native conformations

  • Amphipathic nature complicates crystallization

• Conformational flexibility:

  • The b' subunit likely exhibits significant conformational dynamics

  • This flexibility is essential for function but complicates structural determination

  • Multiple conformational states may exist depending on ATP synthase assembly state

• Interaction-dependent structure:

  • Native structure may depend on interactions with other ATP synthase subunits

  • Isolated b' subunit may not adopt physiologically relevant conformations

  • Co-expression with partner subunits may be necessary for structural studies

• Methodological approaches to overcome these challenges:

  • Nanodiscs or lipid cubic phase crystallization for membrane-associated regions

  • Cryo-electron microscopy for visualization of conformational states

  • Integrated approaches combining NMR for dynamic regions with X-ray crystallography for stable domains

  • Hydrogen-deuterium exchange mass spectrometry to map flexible regions

  • Crosslinking mass spectrometry to capture interaction interfaces

How can researchers investigate the role of ATP synthase in D. shibae's adaptation to anaerobic conditions?

Investigating ATP synthase's role in D. shibae's anaerobic adaptation requires multiple experimental approaches:

• Transcriptional regulation analysis:

  • Perform RNA-Seq comparing aerobic versus anaerobic growth conditions

  • Create reporter gene fusions (atpG-lacZ) to monitor expression under different oxygen levels

  • Analyze potential regulatory sites in the atpG promoter region for binding by FnrL, DnrD, DnrE, or DnrF regulators

• Mutant strain characterization:

  • Generate atpG deletion or point mutants and assess anaerobic growth

  • Construct strains with modified ATP synthase expression levels

  • Examine cross-effects with regulatory mutants (ΔfnrL, ΔdnrD, ΔdnrE, ΔdnrF)

• Biochemical and biophysical analyses:

  • Measure ATP synthesis rates in membrane vesicles from aerobically and anaerobically grown cells

  • Determine P/O ratios (ATP produced per oxygen consumed) under different growth conditions

  • Assess ATP synthase assembly state and subunit stoichiometry using blue native PAGE

• Comparative studies:

  • Analyze ATP synthase activity in D. shibae versus strictly aerobic or anaerobic bacteria

  • Examine post-translational modifications of ATP synthase subunits under different oxygen tensions

  • Compare proton motive force generation across metabolic modes

The anaerobic shift experiments with regulatory mutants provide a useful experimental framework. While ΔdnrD mutants show a clear anaerobic growth defect, ΔfnrL and ΔdnrF mutants surprisingly grow to higher cell densities than wild-type under anaerobic conditions , suggesting complex regulatory interactions affecting energy metabolism.

What purification strategies yield the highest purity and activity of recombinant D. shibae ATP synthase subunit b'?

Optimized purification of recombinant D. shibae ATP synthase subunit b' requires a multi-step approach:

• Membrane extraction optimization:

  • Test multiple detergents (DDM, LMNG, LDAO) for efficient extraction

  • Optimize detergent:protein ratios (typically 2-5:1 w/w)

  • Include stabilizing agents (glycerol 10-20%, specific lipids)

• Affinity chromatography:

  • IMAC (Immobilized Metal Affinity Chromatography) for His-tagged constructs

  • Critical wash steps with low imidazole (20-40 mM) to remove non-specific binding

  • Elution with imidazole gradient (100-300 mM) to separate populations

• Secondary purification steps:

  • Size exclusion chromatography to remove aggregates and isolate homogeneous populations

  • Ion exchange chromatography to separate differently charged species

  • Affinity purification using ATP synthase partner proteins as bait

• Quality control metrics:

  • SDS-PAGE purity >95%

  • Single symmetric peak on size exclusion chromatography

  • Intact mass verification by mass spectrometry

  • Thermal stability assessment by differential scanning fluorimetry

Purification yield comparison table:

How can researchers accurately measure ATP synthase activity in D. shibae membrane preparations?

Accurate measurement of ATP synthase activity in D. shibae membrane preparations requires consideration of the organism's unique metabolic capabilities:

• Membrane preparation methods:

  • Gentle cell disruption using French press or sonication

  • Differential centrifugation to isolate membrane fractions

  • Storage in appropriate buffers with glycerol to maintain stability

• ATP synthesis assays:

  • Luciferin-luciferase based real-time ATP detection

  • Acid-base transition method to generate artificial proton gradient

  • Important controls include oligomycin inhibition to determine ATP synthase-specific activity

• ATP hydrolysis assays:

  • Coupled enzyme assays (pyruvate kinase/lactate dehydrogenase)

  • Malachite green phosphate detection

  • Phenol red-based pH change measurements

• Special considerations for D. shibae:

  • Measure activity under different growth conditions (aerobic, microaerobic, anaerobic with nitrate)

  • Account for light effects given D. shibae's photosynthetic capabilities

  • Compare activity across growth phases to capture regulatory changes

• Data normalization approaches:

  • Protein concentration (Bradford or BCA assay)

  • Bacteriochlorophyll content for photosynthetically grown cells

  • Cytochrome content for respiratory chain normalization

These methodological approaches provide comprehensive assessment of ATP synthase activity within the context of D. shibae's complex energy metabolism.

What experimental approaches can determine the stoichiometry of ATP synthase subunits in D. shibae?

Determining ATP synthase subunit stoichiometry in D. shibae requires multiple complementary approaches:

• Quantitative mass spectrometry:

  • Isotope-labeled reference peptides for absolute quantification (AQUA)

  • Label-free quantification comparing peak intensities

  • TMT or iTRAQ labeling for relative subunit abundance

  • Selected reaction monitoring (SRM) for targeted quantification

• Structural biology approaches:

  • Cryo-electron microscopy of intact ATP synthase complexes

  • Single-particle analysis to determine subunit copy numbers

  • X-ray crystallography of subcomplexes

• Biochemical methods:

  • Blue native PAGE combined with second-dimension SDS-PAGE

  • Densitometric analysis of stained gels

  • Western blotting with subunit-specific antibodies

  • Chemical crosslinking followed by mass spectrometry

• Genetic approaches:

  • Construction of strains with tagged ATP synthase subunits

  • Co-immunoprecipitation combined with quantitative proteomics

  • Sequential affinity purification to isolate intact complexes

These approaches can reveal whether D. shibae ATP synthase follows the typical F₁F₀ bacterial stoichiometry or has adaptations reflecting its unique metabolic capabilities and marine environment.

How should researchers design site-directed mutagenesis experiments to study functional domains of D. shibae ATP synthase subunit b'?

Effective site-directed mutagenesis strategies for D. shibae ATP synthase subunit b' should target key functional domains:

• Target selection based on domain structure:

  • N-terminal membrane-spanning domain: hydrophobic residues for membrane anchoring

  • Central dimerization region: residues involved in b-b' interactions

  • C-terminal domain: residues that interact with F₁ subunits

• Mutation design principles:

  • Conservative substitutions to test specific interactions (e.g., Leu→Ile)

  • Charge reversals to disrupt ionic interactions (e.g., Asp→Lys)

  • Alanine scanning to identify essential residues

  • Introduction or removal of cysteine residues for crosslinking studies

• Experimental validation approaches:

  • In vitro assembly assays with purified components

  • ATP synthesis/hydrolysis activity measurements

  • Complementation of E. coli atpG mutants

  • Growth phenotype analysis under different metabolic conditions

• Integration with structural information:

  • Homology modeling based on related bacterial ATP synthases

  • Molecular dynamics simulations to predict mutation effects

  • Crosslinking mass spectrometry to validate interaction interfaces

When designing mutations, researchers should consider D. shibae's unique metabolic flexibility, which may require ATP synthase adaptations not present in model organisms like E. coli.

How does D. shibae ATP synthase activity integrate with the organism's complex regulatory network for energy metabolism?

D. shibae employs an intricate regulatory network to coordinate its diverse energy metabolism pathways, with ATP synthase functioning at the nexus of these systems:

• FnrL-centered regulatory cascade:

  • The oxygen-sensing regulator FnrL controls expression of high-affinity cbb₃-type cytochrome c oxidase and represses low-affinity aa₃-type cytochrome c oxidase

  • This oxygen-responsive regulation affects electron transport chain composition and subsequently proton motive force generation for ATP synthase

  • The FnrL cascade includes activation of three Dnr regulators (DnrD, DnrE, DnrF) that further modulate energy metabolism genes

• Integration with photosynthetic regulation:

  • Blue light sensing through the LOV-protein LdaP affects photosynthetic gene expression via antirepression of PpsR

  • PpsR normally represses the bchFNBHLM operon for bacteriochlorophyll biosynthesis

  • Photosynthetic activity generates additional proton motive force for ATP synthase

• Coordination with denitrification pathways:

  • Under anaerobic conditions, denitrification becomes a primary energy source

  • The intergenic region between nirS and nosR2 contains Fnr/Dnr binding sites critical for transcriptional activation

  • ATP synthase must adapt to utilize the proton gradient generated during denitrification

• Metabolic state-dependent regulation:

  • Growth phase-dependent changes in gene expression during co-culture with dinoflagellates

  • Quorum sensing systems potentially modulating ATP synthase activity

  • CtrA phosphorelay affecting flagellar biosynthesis and indirectly energy demands

Research at this frontier requires systems biology approaches integrating transcriptomics, proteomics, and metabolomics to map the complex regulatory networks governing ATP synthase activity across different environmental conditions.

What structural adaptations in D. shibae ATP synthase enable function across diverse metabolic modes?

D. shibae's ATP synthase likely contains structural adaptations to support function across aerobic respiration, anaerobic denitrification, and aerobic anoxygenic photosynthesis:

• Catalytic site modifications:

  • Potential amino acid substitutions in β subunit catalytic sites

  • Adaptations that optimize ATP synthesis at varying proton motive force magnitudes

  • Modifications that maintain function despite fluctuating intracellular pH

• Proton channel adaptations:

  • Potential variations in c-ring stoichiometry affecting H⁺/ATP ratio

  • Specialized a-subunit residues optimizing proton flow under variable gradients

  • Structural elements that maintain proton channel integrity during metabolic transitions

• Regulatory domain innovations:

  • Unique interaction sites for metabolic state-sensing regulatory proteins

  • Allosteric regulation mechanisms responsive to energy charge

  • Potential post-translational modification sites for activity modulation

• Stability features:

  • Salt bridges and hydrophobic interactions stabilizing the complex in marine environments

  • Structural elements maintaining assembly during rapid metabolic transitions

  • Interface modifications that optimize interaction with diverse membrane compositions

Research into these structural adaptations requires integrated structural biology approaches combining cryo-EM, X-ray crystallography, mass spectrometry, and functional assays to connect structural features with metabolic versatility.

How do post-translational modifications regulate D. shibae ATP synthase activity under changing environmental conditions?

Post-translational modifications (PTMs) likely play crucial roles in regulating D. shibae ATP synthase activity across environmental transitions:

• Potential modification types:

  • Phosphorylation of regulatory sites in response to energy status

  • Acetylation affecting protein-protein interactions within the complex

  • Redox-sensitive modifications (e.g., disulfide bond formation) responding to oxygen levels

  • Potential lipid modifications anchoring peripheral subunits

• Environmental response patterns:

  • Oxygen-dependent modifications coordinated with the FnrL regulatory cascade

  • Light-responsive modifications linked to photosynthetic activity

  • Salt-dependent modifications reflecting marine environment adaptation

  • Growth phase-specific modifications during symbiotic interactions

• Experimental approaches to investigate PTMs:

  • Phosphoproteomics comparing ATP synthase subunits across growth conditions

  • Targeted mass spectrometry to quantify specific modifications

  • Site-directed mutagenesis of modified residues to assess functional impact

  • In vitro reconstitution with modified and unmodified subunits

• Coordination with regulatory networks:

  • Integration with quorum sensing systems during dinoflagellate interactions

  • Connection to CtrA phosphorelay affecting multiple cellular processes

  • Relationship to stress response pathways during environmental transitions

This research area represents a frontier in understanding how D. shibae rapidly adapts its energy generation systems to environmental changes through dynamic post-translational modification networks.

What are common pitfalls in recombinant expression of D. shibae ATP synthase subunit b' and how can they be addressed?

Recombinant expression of D. shibae ATP synthase subunit b' presents several challenges with specific troubleshooting strategies:

• Expression toxicity:

  • Problem: Growth inhibition following induction

  • Solutions:

    • Use tightly regulated expression systems (e.g., pET with T7 lysozyme)

    • Reduce inducer concentration (0.1-0.5 mM IPTG instead of 1 mM)

    • Lower expression temperature (16-18°C)

    • Use specialized strains (C41(DE3), C43(DE3)) designed for toxic proteins

• Protein aggregation:

  • Problem: Formation of inclusion bodies

  • Solutions:

    • Fusion with solubility tags (MBP, SUMO, TrxA)

    • Co-expression with chaperones (GroEL/ES, DnaK/J)

    • Addition of mild detergents (0.05% DDM) to lysis buffer

    • Extraction and refolding protocols optimized for membrane proteins

• Poor yield:

  • Problem: Low expression levels

  • Solutions:

    • Codon optimization for expression host

    • Optimize growth media (2YT, TB, or auto-induction media)

    • Use fermenter for high-density cultivation

    • Extend expression time at lower temperatures (36-48h at 18°C)

• Proteolytic degradation:

  • Problem: Multiple bands or smearing on SDS-PAGE

  • Solutions:

    • Include multiple protease inhibitors in all buffers

    • Use protease-deficient host strains (BL21, HB2151)

    • Design constructs with improved stability based on secondary structure prediction

    • Maintain samples at 4°C throughout purification

These troubleshooting strategies require systematic optimization for the specific properties of D. shibae ATP synthase subunit b', which may differ from model organism homologs.

How can researchers distinguish between direct and indirect effects when studying ATP synthase function in D. shibae mutants?

Distinguishing direct from indirect effects in ATP synthase studies requires multiple control experiments and complementary approaches:

• Genetic strategies:

  • Use point mutations rather than gene deletions where possible

  • Create partial function alleles that separate different aspects of ATP synthase function

  • Apply inducible or conditional expression systems for temporal control

  • Construct compensatory mutations to test specific interaction models

• Biochemical approaches:

  • Isolate membrane vesicles for direct activity measurements

  • Reconstitute purified components in liposomes

  • Use specific inhibitors (oligomycin, DCCD) with appropriate controls

  • Measure multiple parameters (ATP synthesis, ATP hydrolysis, proton pumping)

• Control experiments for gene regulatory studies:

  • Use reporter gene fusions to monitor expression of multiple genes

  • Perform chromatin immunoprecipitation to confirm direct regulator binding

  • Create mutations in predicted binding sites to verify regulatory mechanisms

  • Complement mutants with wild-type and mutant alleles

• Physiological context controls:

  • Monitor multiple metabolic parameters (oxygen consumption, NAD+/NADH ratio, membrane potential)

  • Compare phenotypes across different growth conditions

  • Measure growth rates, cell morphology, and other physiological parameters

  • Use metabolomics to capture global metabolic effects

These approaches help separate direct effects on ATP synthase from indirect consequences of disrupting cellular energy metabolism.

How might emerging technologies advance our understanding of D. shibae ATP synthase regulation and function?

Several emerging technologies will likely transform research on D. shibae ATP synthase:

• Cryo-electron tomography:

  • Direct visualization of ATP synthase in native membrane environments

  • Mapping spatial distribution and organization within the cell

  • Capturing conformational states under different metabolic conditions

• Single-molecule techniques:

  • Fluorescence resonance energy transfer (FRET) to observe rotational dynamics

  • Magnetic tweezers to measure torque generation

  • Single-molecule force spectroscopy to study subunit interactions

  • High-speed AFM to visualize conformational changes in real-time

• Advanced genetic tools:

  • CRISPR-Cas9 genome editing for precise manipulation of atpG and regulatory elements

  • Inducible CRISPRi for temporal control of gene expression

  • Multiplex genome engineering to study combinatorial effects

  • Site-specific incorporation of unnatural amino acids for specialized probes

• Integrative multi-omics:

  • Combined transcriptomics, proteomics, and metabolomics approaches

  • Flux balance analysis incorporating ATP synthase activity parameters

  • Machine learning models predicting ATP synthase regulation from multi-omics data

  • Spatial transcriptomics and proteomics revealing subcellular heterogeneity

These technologies will enable researchers to address fundamental questions about how D. shibae ATP synthase functions within the context of the organism's complex metabolic networks and environmental adaptations.

What are the implications of D. shibae ATP synthase research for understanding bacterial adaptation to changing marine environments?

Research on D. shibae ATP synthase has broader implications for understanding bacterial adaptation to marine environments:

• Climate change adaptation:

  • Insights into how marine bacteria maintain energy homeostasis despite temperature fluctuations

  • Understanding adaptations to changing oxygen levels in warming oceans

  • Mechanisms for coping with altered photosynthetic conditions due to water clarity changes

• Ecological interactions:

  • Energy economics of symbiotic relationships with marine algae

  • Metabolic transitions underlying mutualistic-to-pathogenic switches

  • ATP synthase modifications that support specialized ecological niches

• Evolutionary adaptations:

  • Comparative analysis with ATP synthases from other marine bacteria

  • Identification of convergent adaptations to marine environments

  • Horizontal gene transfer patterns affecting energy metabolism genes

• Biogeochemical cycling:

  • Role of efficient energy conservation in carbon and nitrogen cycling

  • Impact of bacterial ATP synthase efficiency on marine ecosystem productivity

  • Connections between ATP synthesis and production of climate-active compounds