Deciphering the molecular principles of bacterial metabolosome biogenesis
Lead Research Organisation:
University of Liverpool
Department Name: Institute of Integrative Biology
Abstract
Pathogenic bacteria, such as Salmonella, thrive in mammalian intestines and can cause severe health issues in human, including food poisoning, massive gut inflammation and cardiovascular disease. There were estimated 535,000 cases of Salmonella gastrointestinal infections worldwide in 2017, and 91,857 cases in the EU in 2018. Salmonella cells produce a specialised nano-scale organelle, known as the bacterial microcompartment. These organelles provide a suite of unique metabolic advantages that allow Salmonella to become the predominant species in the hostile environment of the host gut. The organelle uses a shell that is made of thousands of proteins to sequester multiple enzymes used for 1,2-propanediol utilisation (Pdu). This unique structure allows the Pdu organelles to protect bacterial cells from toxic metabolites and to enhance the cell's metabolism. Although the importance of Pdu organelles for the metabolism of bacterial pathogens is appreciated, little is known about how bacterial cells generate and then modulate these organelles to confer adaptive cellular metabolism to survive in the sophisticated gut environment.
We have recently reported the exact protein stoichiometry of Pdu organelles and have established a new structural model of the organelle. We have also developed systems for tagging Pdu proteins with fluorescent markers and depleting target proteins, so that we can track specific building proteins using microscopes and study their functions in bacterial cells. Using the developed systems, we have discovered that the cargo enzymes and shell proteins self-assemble independently in Salmonella. We have also shown that the locations and movement of Pdu organelles are confined within the bacterial cell. Standing on these exciting scientific and technical breakthroughs and an established research team with complementary expertise, we now aim to do an in-depth exploration of how Pdu organelles are synthesized and how the organisation of Pdu organelles is coordinated within the Salmonella cell.
We will first determine the multi-step assembly that individual building proteins undergo to form higher-ordered assemblies, identify the proteins that make up the enzyme and shell assemblies, and elucidate how enzyme and shell assemblies associate together to form an intact organelle. In the second section of our programme, we will characterise the structures and functions of small linker proteins that drive the assembly of cargos to have a "liquid-like" dynamic phase and ascertain that the phase separation mechanism is vital for mediating the protein interactions and formation of a functional protein organelle. Finally, we will use state-of-the-art fluorescence microscopy to probe how the Pdu organelles are generated, located, and modulated to perform such important functions in bacterial cells.
This ambitious and multidisciplinary research project has both fundamental and applied significance. Pdu MCPs represents an ideal model system for uncovering the principles of protein self-assembly and the generation of multi-protein complexes in biology. We will learn the basic physics and chemistry of how thousands of proteins assemble together to build a functional entity within a bacterial cell, and determine how the cell precisely and efficiently controls the formation and function of metabolic organelles. We anticipate that our findings will provide a deeper understanding of the biosynthesis and maintenance of natural bacterial organelles and protein assemblies. The research may inform strategies for the engineering of biological "factories" for the enhancement of cell metabolism and energy production in diverse biotechnological applications. Moreover, the essential protein-protein interactions that we find are required to mediate the assembly of Pdu organelles could represent novel therapeutic targets to disrupt the production of Pdu organelles and thus ablate the ability of Salmonella to thrive in the human gut.
We have recently reported the exact protein stoichiometry of Pdu organelles and have established a new structural model of the organelle. We have also developed systems for tagging Pdu proteins with fluorescent markers and depleting target proteins, so that we can track specific building proteins using microscopes and study their functions in bacterial cells. Using the developed systems, we have discovered that the cargo enzymes and shell proteins self-assemble independently in Salmonella. We have also shown that the locations and movement of Pdu organelles are confined within the bacterial cell. Standing on these exciting scientific and technical breakthroughs and an established research team with complementary expertise, we now aim to do an in-depth exploration of how Pdu organelles are synthesized and how the organisation of Pdu organelles is coordinated within the Salmonella cell.
We will first determine the multi-step assembly that individual building proteins undergo to form higher-ordered assemblies, identify the proteins that make up the enzyme and shell assemblies, and elucidate how enzyme and shell assemblies associate together to form an intact organelle. In the second section of our programme, we will characterise the structures and functions of small linker proteins that drive the assembly of cargos to have a "liquid-like" dynamic phase and ascertain that the phase separation mechanism is vital for mediating the protein interactions and formation of a functional protein organelle. Finally, we will use state-of-the-art fluorescence microscopy to probe how the Pdu organelles are generated, located, and modulated to perform such important functions in bacterial cells.
This ambitious and multidisciplinary research project has both fundamental and applied significance. Pdu MCPs represents an ideal model system for uncovering the principles of protein self-assembly and the generation of multi-protein complexes in biology. We will learn the basic physics and chemistry of how thousands of proteins assemble together to build a functional entity within a bacterial cell, and determine how the cell precisely and efficiently controls the formation and function of metabolic organelles. We anticipate that our findings will provide a deeper understanding of the biosynthesis and maintenance of natural bacterial organelles and protein assemblies. The research may inform strategies for the engineering of biological "factories" for the enhancement of cell metabolism and energy production in diverse biotechnological applications. Moreover, the essential protein-protein interactions that we find are required to mediate the assembly of Pdu organelles could represent novel therapeutic targets to disrupt the production of Pdu organelles and thus ablate the ability of Salmonella to thrive in the human gut.
Technical Summary
Bacterial microcompartments (BMCs) are intracellular proteinaceous organelles that spatially organise and confine metabolic reactions. The 1,2-propanediol utilisation microcompartments (Pdu MCPs) in pathogenic Salmonella sequester enzymatic pathways that produce toxic metabolites using a virus-like shell. This confers growth advantages to Salmonella within the human microbiome. The pathway that thousands of protein subunits self-assemble in order and time to form functional Pdu MCPs in cells remains elusive. We have recently developed approaches for the isolation, proteomics, genetic modification and cell imaging to explore the Pdu MCP biogenesis in Salmonella. Our preliminary results indicated that Pdu MCP shell and core enzymes undergo the separated assembly and that formation of enzyme assemblies is mediated by specific intrinsically disordered short peptides.
We hypothesise that Pdu MCPs possess a "concomitant" biogenesis pathway and cargo enzymes form liquid-like protein assemblies driven by liquid-liquid phase separation. Using proteomics and confocal microscopy, we will first explore how individual building components assemble to form the shell and enzyme assemblage and then the entire Pdu MCPs. Next, we will determine the structures of key disordered peptides and their interactions with cargos using NMR and isothermal titration calorimetry. We will also corroborate the liquid-like properties of enzyme assemblies using fluorescence recovery after photobleaching. Finally, we will use live-cell fluorescence imaging to explore the biogenic sites, subcellular partitioning and movement of Pdu MCPs, to elucidate how Salmonella integrates Pdu MCPs with the bacterial cytoskeleton. Advanced knowledge of Pdu MCP biogenesis, protein interactions and encapsulation will inform the engineering of bio-factories for optimising metabolic performance, producing toxic proteins, and may lead to therapeutics for preventing colonisation of the human GI tract by Salmonella.
We hypothesise that Pdu MCPs possess a "concomitant" biogenesis pathway and cargo enzymes form liquid-like protein assemblies driven by liquid-liquid phase separation. Using proteomics and confocal microscopy, we will first explore how individual building components assemble to form the shell and enzyme assemblage and then the entire Pdu MCPs. Next, we will determine the structures of key disordered peptides and their interactions with cargos using NMR and isothermal titration calorimetry. We will also corroborate the liquid-like properties of enzyme assemblies using fluorescence recovery after photobleaching. Finally, we will use live-cell fluorescence imaging to explore the biogenic sites, subcellular partitioning and movement of Pdu MCPs, to elucidate how Salmonella integrates Pdu MCPs with the bacterial cytoskeleton. Advanced knowledge of Pdu MCP biogenesis, protein interactions and encapsulation will inform the engineering of bio-factories for optimising metabolic performance, producing toxic proteins, and may lead to therapeutics for preventing colonisation of the human GI tract by Salmonella.
Organisations
- University of Liverpool (Lead Research Organisation)
- Chinese Academy of Sciences (Collaboration)
- Lancaster University (Collaboration)
- Lawrence Berkeley National Laboratory (Collaboration)
- Australian National University (ANU) (Collaboration)
- UNIVERSITY OF KENT (Collaboration)
- Huazhong Agricultural University (Collaboration)
- UNIVERSITY OF LIVERPOOL (Collaboration)
- DIAMOND LIGHT SOURCE (Collaboration)
- IMPERIAL COLLEGE LONDON (Collaboration)
- University of Science and Technology of China USTC (Collaboration)
- Autonomous University of Madrid (Collaboration)
People |
ORCID iD |
Luning Liu (Principal Investigator) |
Publications
Bracun L
(2023)
Cryo-EM structure of a monomeric RC-LH1-PufX supercomplex with high-carotenoid content from Rhodobacter capsulatus
in Structure
Bracun L
(2021)
Cryo-EM structure of the photosynthetic RC-LH1-PufX supercomplex at 2.8-Å resolution.
in Science advances
Cao P
(2022)
Structural basis for the assembly and quinone transport mechanisms of the dimeric photosynthetic RC-LH1 supercomplex.
in Nature communications
Chen T
(2023)
Producing fast and active Rubisco in tobacco to enhance photosynthesis.
in The Plant cell
Chen T
(2023)
Engineering a-carboxysomes into plant chloroplasts to support autotrophic photosynthesis.
in Nature communications
Chen T
(2022)
Incorporation of Functional Rubisco Activases into Engineered Carboxysomes to Enhance Carbon Fixation.
in ACS synthetic biology
Evans SL
(2023)
Single-particle cryo-EM analysis of the shell architecture and internal organization of an intact a-carboxysome.
in Structure (London, England : 1993)
Fang S
(2021)
Molecular mechanism underlying transport and allosteric inhibition of bicarbonate transporter SbtA.
in Proceedings of the National Academy of Sciences of the United States of America
Faulkner M
(2023)
Chemoautotrophic production of gaseous hydrocarbons, bioplastics and osmolytes by a novel Halomonas species.
in Biotechnology for biofuels and bioproducts
Huang J
(2022)
Probing the Internal pH and Permeability of a Carboxysome Shell
in Biomacromolecules
Title | Additional file 2 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 2: Table 1. Proteins/peptides used in ALACAT B. Figure S1. Extracted ion chromatogram and peptide coverage map for short ALACAT 301. Figure S2. Extracted ion chromatogram and peptide coverage map for short ALACAT 302. Figure S3. Extracted ion chromatogram and peptide coverage map for short ALACAT 301. Figure S4. Extracted ion chromatogram and peptide coverage map for short ALACAT 304. Figure S5. Extracted ion chromatogram and peptide coverage map for short ALACAT 305. Figure S6. Extracted ion chromatogram and peptide coverage map for long ALACAT B. Figure S7. Wheat germ proteins present in purified QconCATs. Figure S8. Uncropped SDS-PAGE images. |
Type Of Art | Film/Video/Animation |
Year Produced | 2021 |
URL | https://springernature.figshare.com/articles/presentation/Additional_file_2_of_Construction_of_la_ca... |
Title | Additional file 2 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 2: Table 1. Proteins/peptides used in ALACAT B. Figure S1. Extracted ion chromatogram and peptide coverage map for short ALACAT 301. Figure S2. Extracted ion chromatogram and peptide coverage map for short ALACAT 302. Figure S3. Extracted ion chromatogram and peptide coverage map for short ALACAT 301. Figure S4. Extracted ion chromatogram and peptide coverage map for short ALACAT 304. Figure S5. Extracted ion chromatogram and peptide coverage map for short ALACAT 305. Figure S6. Extracted ion chromatogram and peptide coverage map for long ALACAT B. Figure S7. Wheat germ proteins present in purified QconCATs. Figure S8. Uncropped SDS-PAGE images. |
Type Of Art | Film/Video/Animation |
Year Produced | 2021 |
URL | https://springernature.figshare.com/articles/presentation/Additional_file_2_of_Construction_of_la_ca... |
Description | We have successfully elucidated the biogenesis pathway of microcompartments in pathogenic bacteria and developed bioimaging techniques for live-cell imaging for physiological determination. The research findings have led to a paper in Nature Communications (Nature Commun, 2022, 13: 2920). |
Exploitation Route | publications, techniques |
Sectors | Healthcare Manufacturing including Industrial Biotechology Pharmaceuticals and Medical Biotechnology |
Description | By gaining a mechanistic understanding of bacterial organelles, our study paves the way for engineering BMCs to enhance enzymatic activities and encapsulate toxic metabolites, and offers new therapeutic targets for Salmonella infection in the gut. |
First Year Of Impact | 2023 |
Sector | Chemicals,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology |
Impact Types | Societal Economic |
Description | Engineering functional CO2-fixing organelles to enhance plant photosynthesis |
Amount | £208,358 (GBP) |
Funding ID | RPG-2021-286 |
Organisation | The Leverhulme Trust |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 09/2021 |
End | 09/2024 |
Title | AFM |
Description | atomic force microscopy imaging on biological samples |
Type Of Material | Technology assay or reagent |
Year Produced | 2018 |
Provided To Others? | Yes |
Impact | We have applied high-resolution AFM imaging on many biological samples. Recently we have established a hybrid AFM/confocal/TIRF microscopy for studying cell dynamics. Structural variability, coordination, and adaptation of a native photosynthetic machinery. Zhao LS, Huokko T, Wilson S, Simpson DM, Wang Q, Ruban AV, Mullineaux CW, Zhang YZ*, Lu-Ning Liu*. Nature Plants, 2020, 6(7): 869-882. DOI: 10.1038/s41477-020-0694-3. AFM imaging on amyloid morphology provides further motivation to investigate the role of oxidative stress in AMA pathogenicity. The study has led to a paper published: Davies HA, Phelan MM, Wilkinson MC, Migrino RQ, Truran S, Franco DA, Liu LN, Longmore CJ, Madine J. Oxidative stress alters morphology and toxicity of aortic medial amyloid. Biophys J, 2015, 109(11): 2363-2370. Unfolding pathway and intermolecular interactions of the cytochrome subunit in the bacterial photosynthetic reaction center. Leanne C. Miller, Longsheng Zhao, Daniel P. Canniffe, David Martin, Lu-Ning Liu*. Biochim Biophys Acta - Bioenergetics, 2020, 1861(8): 148204, DOI: 10.1016/j.bbabio.2020.148204. AFM imaging on exosome structure has revealed that CLL cells secrete exosomes that alter the transcriptome and behaviour of recipient cells. Such communication with microenvironment is likely to have an important role in CLL disease biology. The study has led to a paper published: Farahani M, Rubbi C, Liu LN, Slupsky JR, Kalakonda N. CLL exosomes modulate the transcriptome and behaviour of recipient stromal cells and are selectively enriched in miR-202-3p. PLoS ONE, 2015, 10(10): e0141429. |
Title | AFM force measurement to study protein interaction |
Description | apply atomic force microscopy force measurement to study the protein unfolding and interaction of membrane protein complexes |
Type Of Material | Technology assay or reagent |
Year Produced | 2020 |
Provided To Others? | Yes |
Impact | Using state-of-the-art Atomic Force Microscopy (AFM), we have deciphered the nanoscale structure of the photosynthetic membranes that are extracted from a purple photosynthetic bacterium (Blastochloris viridis). Additionally, we applied single-molecule force spectroscopy (SMFS) to "pull" out protein peptides from the photosynthetic complexes in their working conditions. This allowed us to monitor the stepwise unfolding process of the structural components of photosynthetic complexes and detect the mechanical forces required in the unfolding process. Unfolding pathway and intermolecular interactions of the cytochrome subunit in the bacterial photosynthetic reaction center. Leanne C. Miller, Longsheng Zhao, Daniel P. Canniffe, David Martin, Lu-Ning Liu*. Biochim Biophys Acta - Bioenergetics, 2020, 1861(8): 148204, |
URL | https://www.sciencedirect.com/science/article/pii/S0005272820300542?via%3Dihub |
Title | Absolute proteomic quantification of protein assemblies |
Description | use Qconcat method based on quantitative mass spectrometry to determine the stoichiometry of protein components in macromolecular complexes. |
Type Of Material | Technology assay or reagent |
Year Produced | 2019 |
Provided To Others? | No |
Impact | Decoding the stoichiometric composition and organisation of bacterial metabolosomes. Yang M, Simpson DM, Wenner N, Brownridge P, Harman VM, Hinton JCD, Beynon RJ, Lu-Ning Liu* Nature Communications, 2020, 11(1): 1976. DOI: 10.1038/s41467-020-15888-4. |
URL | https://www.nature.com/articles/s41467-020-15888-4 |
Title | Additional file 1 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 1. DNA and protein sequences of construct, translated sequences and cloning syntaxes. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_1_of_Construction_of_la_carte_Q... |
Title | Additional file 1 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 1. DNA and protein sequences of construct, translated sequences and cloning syntaxes. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_1_of_Construction_of_la_carte_Q... |
Title | Additional file 3 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 3. DNA products from long combinatorial study. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_3_of_Construction_of_la_carte_Q... |
Title | Additional file 3 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 3. DNA products from long combinatorial study. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_3_of_Construction_of_la_carte_Q... |
Title | Additional file 4 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 4. DNA products from long combinatorial study. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_4_of_Construction_of_la_carte_Q... |
Title | Additional file 4 of Construction of à la carte QconCAT protein standards for multiplexed quantification of user-specified target proteins |
Description | Additional file 4. DNA products from long combinatorial study. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://springernature.figshare.com/articles/dataset/Additional_file_4_of_Construction_of_la_carte_Q... |
Title | Supplementary File 1 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_File_1_from_Characterizing_the_supercomplex_a... |
Title | Supplementary File 1 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_File_1_from_Characterizing_the_supercomplex_a... |
Title | Supplementary File 2 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_File_2_from_Characterizing_the_supercomplex_a... |
Title | Supplementary File 2 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_File_2_from_Characterizing_the_supercomplex_a... |
Title | Supplementary Table 1 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_Table_1_from_Characterizing_the_supercomplex_... |
Title | Supplementary Table 1 from Characterizing the supercomplex association of photosynthetic complexes in cyanobacteria |
Description | The light reactions of photosynthesis occur in thylakoid membranes that are densely packed with a series of photosynthetic complexes. The lateral organization and close association of photosynthetic complexes in native thylakoid membranes are vital for efficient light harvesting and energy transduction. Recently, analysis of the interconnections between photosynthetic complexes to form supercomplexes has garnered great interest. In this work, we report a method integrating immunoprecipitation, mass spectrometry and atomic force microscopy to identify the inter-complex associations of photosynthetic complexes in thylakoid membranes from the cyanobacterium Synechococcus elongatus PCC 7942. We characterize the preferable associations between individual photosynthetic complexes and binding proteins involved in the complex-complex interfaces, permitting us to propose the structural models of photosynthetic complex associations that promote the formation of photosynthetic supercomplexes. We also identified other potential binding proteins with the photosynthetic complexes, suggesting the highly connecting networks associated with thylakoid membranes. This study provides mechanistic insight into the physical interconnections of photosynthetic complexes and potential partners, which are crucial for efficient energy transfer and physiological acclimatization of the photosynthetic apparatus. Advanced knowledge of the protein organization and interplay of the photosynthetic machinery will inform rational design and engineering of artificial photosynthetic systems to supercharge energy production. |
Type Of Material | Database/Collection of data |
Year Produced | 2021 |
Provided To Others? | Yes |
URL | https://rs.figshare.com/articles/dataset/Supplementary_Table_1_from_Characterizing_the_supercomplex_... |
Title | alpha-carboxysome QconCAT quantification |
Description | The entire Skyline project and raw data for a-carboxysome QconCAT quantification have been deposited at Panorama Public with the access URL (https://panoramaweb.org/Wb6olk.url) and the ProteomeXchange ID PXD031494. Raw LC-MSMS data for label-free quantification have been deposited to the ProteomeXchange Consortium via the PRIDE (73) partner repository with the access URL (https://www.ebi.ac.uk/pride/archive/projects/PXD031420). |
Type Of Material | Database/Collection of data |
Year Produced | 2022 |
Provided To Others? | Yes |
Impact | Carboxysomes are anabolic bacterial microcompartments that play an essential role in carbon fixation in cyanobacteria and some chemoautotrophs. This self-assembling organelle encapsulates the key CO2-fixing enzymes, Rubisco, and carbonic anhydrase using a polyhedral protein shell that is constructed by hundreds of shell protein paralogs. The a-carboxysome from the chemoautotroph Halothiobacillus neapolitanus serves as a model system in fundamental studies and synthetic engineering of carboxysomes. Here we adopt a QconCAT-based quantitative mass spectrometry approach to determine the stoichiometric composition of native a-carboxysomes from H. neapolitanus. We further performed an in-depth comparison of the protein stoichiometry of native and recombinant a-carboxysomes heterologously generated in Escherichia coli to evaluate the structural variability and remodeling of a-carboxysomes. Our results provide insight into the molecular principles that mediate carboxysome assembly, which may aid in rational design and reprogramming of carboxysomes in new contexts for biotechnological applications. |
URL | https://www.ebi.ac.uk/pride/archive/projects/PXD031420 |
Description | BMC - De Pablo |
Organisation | Autonomous University of Madrid |
Country | Spain |
Sector | Academic/University |
PI Contribution | BMC isolation and expression, data analysis |
Collaborator Contribution | provide AFM training and imaging analysis of BMC samples |
Impact | secured a Johnston Fund (University of Liverpool, £2155) to support postdoc researcher to visit de Pablo Lab in March 2018. |
Start Year | 2017 |
Description | BMC - Kerfeld |
Organisation | Lawrence Berkeley National Laboratory |
Country | United States |
Sector | Public |
PI Contribution | microscopy imaging of bacterial microcompartment assembly |
Collaborator Contribution | generating constructs and structural analysis |
Impact | One publication in Nano Letters, featured as a cover article Visualization of bacterial microcompartment facet assembly using high-speed atomic force microscopy. M. Sutter, M. Faulkner, C. Aussignargues, B.C. Paasch, S. Barrett, C.A. Kerfeld, L.N. Liu. Nano Letters, 2016, 2016, 16(3): 1590-1595, DOI: 10.1021/acs.nanolett.5b04259. |
Start Year | 2015 |
Description | BMC - Lin |
Organisation | Huazhong Agricultural University |
Country | China |
Sector | Academic/University |
PI Contribution | making genetic constructs and protein purification |
Collaborator Contribution | providing technical support for plant engineering |
Impact | The collaboration has secured a Royal Society Challenge Grant (Enhancing crop photosynthesis and productivity by engineering CO2-fixing organelles. 2017-2018, CH160004. £120K. PI) |
Start Year | 2016 |
Description | BMC - Nixon |
Organisation | Imperial College London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | making cyanobacterial constructs and thylakoid membrane samples |
Collaborator Contribution | plant engineering, biochemical investigation of thylakoid membrane composition. |
Impact | the preliminary collaborative work has helped to secure a BBSRC grant (BB/R003890/1) |
Start Year | 2015 |
Description | BMC - Parry |
Organisation | Lancaster University |
Department | Lancaster Environment Centre |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | build genetic constructs to engineer carboxysomes in plants |
Collaborator Contribution | conduct plant transformation |
Impact | A new collaboration. No outputs yet. This collaboration is multi-disciplinary, including microbiology, biochemistry, synthetic biology and plant engineering. The collaboration has secured a Royal Society Challenge Grant (Enhancing crop photosynthesis and productivity by engineering CO2-fixing organelles. 2017-2018, CH160004. £120K. PI) |
Start Year | 2017 |
Description | BMC - Price |
Organisation | Australian National University (ANU) |
Country | Australia |
Sector | Academic/University |
PI Contribution | microscopy imaging of bacterial microscopy proteins and assemblies |
Collaborator Contribution | generating mutants and sharing protocols |
Impact | Royal Society International Exchange grant: £12,000. A manuscript is in preparation |
Start Year | 2015 |
Description | BMC - Zhou |
Organisation | University of Science and Technology of China USTC |
Country | China |
Sector | Academic/University |
PI Contribution | prepare constructs and proteins |
Collaborator Contribution | structural analysis |
Impact | A manuscript based on the collaborative data is in preparation |
Start Year | 2016 |
Description | Ci transporter - Zhang |
Organisation | Chinese Academy of Sciences |
Department | Shanghai Institute of Plant Physiology and Ecology |
Country | China |
Sector | Academic/University |
PI Contribution | study the physiological functions and localisation of bicarbonate transporters in cyanobacteria |
Collaborator Contribution | study the crystal structures of bicarbonate transporters using X-ray crystallography |
Impact | One manuscript is under review. (Wang C, Sun B, Zhang M, Huang X, Guo H, Chen X, Huang F, Chen T, Mi H, Liu LN, Zhang P (2018) Structural mechanism of the active bicarbonate transporter from cyanobacteria. Sci Adv, under review) |
Start Year | 2017 |
Description | Computational simulations - Jun Gao |
Organisation | Huazhong Agricultural University |
Country | China |
Sector | Academic/University |
PI Contribution | protein structural analysis |
Collaborator Contribution | Computational simulations on protein interactions and electron transfer |
Impact | Million-atom molecular dynamics simulations reveal the interfacial interactions and assembly of plant PSII-LHCII supercomplex. Ruichao Mao, Han Zhang, Lihua Bie, Lu-Ning Liu*, Jun Gao*. RSC Advances, 2023, 13(10): 6699-6712. DOI: 10.1039/d2ra08240c. Multidisciplinary: computational simulations, structural biology, biochemistry, microbiology |
Start Year | 2020 |
Description | Cryo-ET - Zhang |
Organisation | Diamond Light Source |
Country | United Kingdom |
Sector | Private |
PI Contribution | grow cyanobacterial cells in different physiological conditions to regulate thylakoid membrane biogenesis |
Collaborator Contribution | use cryo-electron tomography to study the thylakoid membrane organisation in cyanobacterial cells |
Impact | one collaborative manuscript is in preparation |
Start Year | 2019 |
Description | NMR protein analysis -Lu-Yun Lian |
Organisation | University of Liverpool |
Department | Institute of Integrative Biology |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | Pdu microcompartment protein purification and interaction |
Collaborator Contribution | protein purification and interaction analysis using NMR |
Impact | a manuscript is in preparation |
Start Year | 2017 |
Description | Pdu bioengineering - Warren |
Organisation | University of Kent |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | provide analytical tools and constructs |
Collaborator Contribution | provide plasmids and protocols |
Impact | Not yet |
Start Year | 2021 |
Description | Salmonella physiology -Jay Hinton |
Organisation | University of Liverpool |
Department | Institute of Integrative Biology |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | study Pdu microcompartment biogenesis |
Collaborator Contribution | support Salmonella physiology, genetics and growth assays |
Impact | Biogenesis of a bacterial metabolosome for propanediol utilization. Mengru Yang, Nicolas Wenner, Gregory Dykes, Yan Li, Xiaojun Zhu, Yaqi Sun, Fang Huang, Jay Hinton, Lu-Ning Liu*. Nature Communications, 2022, 13: 2920. DOI: 10.1038/s41467-022-30608-w. It is multi-disciplinary: molecular genetics, microbiology, cell physiology, microscopic imaging, bioinformatics Decoding the stoichiometric composition and organisation of bacterial metabolosomes. Yang M, Simpson DM, Wenner N, Brownridge P, Harman VM, Hinton JCD, Beynon RJ, Lu-Ning Liu*. Nature Communications, 2020, 11(1): 1976. DOI: 10.1038/s41467-020-15888-4. It is multi-disciplinary: molecular genetics, microbiology, biochemistry, microscopic imaging, proteomics |
Start Year | 2017 |
Title | Bacterial microcompartmental shells |
Description | The present invention relates to new synthetically generated carboxysome shells, synthetic shells encapsulating selected catalytically active moieties, cells including the synthetic shells and methods of making the synthetic shells and methods of encapsulation. The invention includes inter alia, uses of the shells as nanoreactors for enhanced catalytic performance of the encapsulated enzymes. |
IP Reference | GB2015810.1 |
Protection | Patent granted |
Year Protection Granted | 2020 |
Licensed | Yes |
Impact | develop a novel technology to create shell structures for diverse biotechnological applications |
Description | Invited talk, DiscoverBMB 2023, annual meeting of the American Society for Biochemistry and Molecular Biology (ASBMB), Mar 25-28, 2023, Seattle, US |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | With a mission to share the latest, most impactful research findings in the molecular life sciences, #DiscoverBMB offers an exciting agenda that includes talks by the field's foremost experts, interactive workshops on the latest trends, technologies and techniques, and an invigorating exhibition of posters, services and products. |
Year(s) Of Engagement Activity | 2023 |
Description | Invited talk, ERC Synergy Symposium |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Invited talk, ERC Synergy Symposium |
Year(s) Of Engagement Activity | 2023 |