Professor Johnjoe McFadden
Professor of Molecular Genetics
Qualifications: BSc (Biochemistry), PhD (Biochemistry)
Email: j.mcfadden@surrey.ac.uk
Phone: Work: 01483 68 6494
Room no: 07 AX 01
Further information
Biography
2002 – present, . Professor of Molecular Genetics, University of Surrey
1994 – 2001 Reader, School of Biological Sciences, University of Surrey.
1988-94 Lecturer, School of Biological Sciences, University of Surrey.
1984-88 Research Fellow. Department of Surgery, St. George's Hospital Medical School, London.
1982-84 Research Fellow, Department of Biochemistry, St. Mary's Hospital Medical School, London.
1982: PhD (Biochemistry) Imperial College, University of London
1978 BSc (Biochemistry) Bedford College, University of London
Research Interests
Systems Biology
Mycobacterial genetics
Pathogenicity of tuberculosis
Neisserial genetics
Pathogenicity of meningococcal meningitis
Bionanotechnology
Research Collaborations
CARBIO network: Marie Curie Research Training Network - Multifunctional Carbon Nanotubes for Biomedical Applications
Publications
Highlights
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(2011) '¹³C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation.'. PLoS PLoS Pathog, United States: 7 (7)Full text is available at: http://epubs.surrey.ac.uk/184899/
Abstract
Mycobacterium tuberculosis requires the enzyme isocitrate lyase (ICL) for growth and virulence in vivo. The demonstration that M. tuberculosis also requires ICL for survival during nutrient starvation and has a role during steady state growth in a glycerol limited chemostat indicates a function for this enzyme which extends beyond fat metabolism. As isocitrate lyase is a potential drug target elucidating the role of this enzyme is of importance; however, the role of isocitrate lyase has never been investigated at the level of in vivo fluxes. Here we show that deletion of one of the two icl genes impairs the replication of Mycobacterium bovis BCG at slow growth rate in a carbon limited chemostat. In order to further understand the role of isocitrate lyase in the central metabolism of mycobacteria the effect of growth rate on the in vivo fluxes was studied for the first time using ¹³C-metabolic flux analysis (MFA). Tracer experiments were performed with steady state chemostat cultures of BCG or M. tuberculosis supplied with ¹³C labeled glycerol or sodium bicarbonate. Through measurements of the ¹³C isotopomer labeling patterns in protein-derived amino acids and enzymatic activity assays we have identified the activity of a novel pathway for pyruvate dissimilation. We named this the GAS pathway because it utilizes the Glyoxylate shunt and Anapleurotic reactions for oxidation of pyruvate, and Succinyl CoA synthetase for the generation of succinyl CoA combined with a very low flux through the succinate--oxaloacetate segment of the tricarboxylic acid cycle. We confirm that M. tuberculosis can fix carbon from CO₂ into biomass. As the human host is abundant in CO₂ this finding requires further investigation in vivo as CO₂ fixation may provide a point of vulnerability that could be targeted with novel drugs. This study also provides a platform for further studies into the metabolism of M. tuberculosis using ¹³C-MFA.
Journal articles
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(2012) 'Drug loading, dispersion stability, and therapeutic efficacy in targeted drug delivery with carbon nanotubes'. Elsevier Carbon, 50 (2), pp. 622-632.Full text is available at: http://epubs.surrey.ac.uk/735956/
Abstract
We have designed a drug delivery system for the anti-cancer drugs doxorubicin and mitoxantrone based on carbon nanotubes, which is stable under biological conditions, allows for sustained release, and promotes selectivity through an active targeting scheme. Carbon nanotubes are particularly promising for this area of application due to their high surface area, allowing for high drug loading, and their unique interaction with cellular membranes. We have taken a systematic approach to PEG conjugation in order to create a formulation of stable and therapeutically effective CNTs. The presented drug delivery system may be a means of improving cancer treatment modalities by reducing drug-related side effects.
- . (2012) 'Protamine and chloroquine enhance gene delivery and expression mediated by RNA-wrapped single walled carbon nanotubes'. Journal of Nanoscience and Nanotechnology, 12 (3), pp. 1739-1747.
- . (2012) 'Rational design of diagnostic and vaccination strategies for tuberculosis.'. Braz J Infect Dis, Brazil: 16 (1), pp. 68-73.
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(2012) 'Cellular localization, accumulation and trafficking of double-walled carbon nanotubes in human prostate cancer cells'. Nano Research, 5 (4), pp. 223-234.Full text is available at: http://epubs.surrey.ac.uk/711610/
Abstract
Carbon nanotubes (CNTs) are at present being considered as potential nanovectors with the ability to deliver therapeutic cargoes into living cells. Previous studies established the ability of CNTs to enter cells and their therapeutic utility, but an appreciation of global intracellular trafficking associated with their cellular distribution has yet to be described. Despite the many aspects of the uptake mechanism of CNTs being studied, only a few studies have investigated internalization and fate of CNTs inside cells in detail. In the present study, intracellular localization and trafficking of RNA-wrapped, oxidized double-walled CNTs (oxDWNT-RNA) is presented. Fixed cells, previously exposed to oxDWNT-RNA, were subjected to immunocytochemical analysis using antibodies specific to proteins implicated in endocytosis; moreover cell compartment markers and pharmacological inhibitory conditions were also employed in this study. Our results revealed that an endocytic pathway is involved in the internalization of oxDWNT-RNA. The nanotubes were found in clathrin-coated vesicles, after which they appear to be sorted in early endosomes, followed by vesicular maturation, become located in lysosomes. Furthermore, we observed co-localization of oxDWNT-RNA with the small GTP-binding protein (Rab 11), involved in their recycling back to the plasma membrane via endosomes from the trans-golgi network. © 2012 Tsinghua University Press and Springer-Verlag Berlin Heidelberg.
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(2012) 'Modeling the binding of peptides on carbon nanotubes and their use as protein and DNA carriers'. Journal of Nanoparticle Research, 14 (2)Full text is available at: http://epubs.surrey.ac.uk/736295/
Abstract
An in-depth study of a novel functionalization of carbon nanotubes for their application as protein and DNA carriers is presented. First, the optimum conditions for the dispersion of singlewalled carbon nanotubes (SWCNTs) with amphiphilic polypeptides were obtained, and the SWCNT-polypeptide complexes were characterized by different techniques (UV-Vis-NIR, CD, and AFM). Based on the properties of the SWCNT-polypeptide complexes, a model that characterizes the adsorption of natural proteins onto SWCNT was described for the first time. This model predicts the adsorption of natural proteins on SWCNTs based on the protein structure and composition, and therefore, allows the design of methods for the preparation of SWCNT-protein complexes. Besides, the use of cationic-designed amphiphilic polypeptides to disperse SWCNTs is applied for subsequent and efficient binding of DNA to carbon nanotubes by a bilayer approach. Therefore, in this article, we develop procedures for the use of SWCNTs as protein and DNA carriers. The systems were delivered into cells showing that the efficiency of delivery is affected by the charge of the complexes, which has important implications in the use of SWCNT as platforms for protein and DNA binding and subsequent use as delivery systems. © Springer Science+Business Media B.V. 2012.
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(2011) 'Interrogation of global mutagenesis data with a genome scale model of Neisseria meningitidis to assess gene fitness in vitro and in sera.'. BioMed Central Ltd Genome Biol, 12 (12)Full text is available at: http://epubs.surrey.ac.uk/629934/
Abstract
BACKGROUND: Neisseria meningitidis is an important human commensal and pathogen that causes several thousand deaths each year, mostly in young children. How the pathogen replicates and causes disease in the host is largely unknown, particularly the role of metabolism in colonization and disease. Completed genome sequences are available for several strains but our understanding of how these data relate to phenotype remains limited. RESULTS: To investigate the metabolism of N. meningitidis we generated and selected a representative Tn5 library on rich medium, a minimal defined medium and in human serum to identify genes essential for growth under these conditions. To relate these data to a systems-wide understanding of the pathogen's biology we constructed a genome-scale metabolic network: Nmb_iTM560. This model was able to distinguish essential and non-essential genes as predicted by the global mutagenesis. These essentiality data, the library and the Nmb_iTM560 model are powerful and widely applicable resources for the study of meningococcal metabolism and physiology. We demonstrate the utility of these resources by predicting and demonstrating metabolic requirements on minimal medium such as a requirement for PEP carboxylase, and by describing the nutritional and biochemical status of N. meningitidis when grown in serum, including a requirement for both the synthesis and transport of amino acids. CONCLUSIONS: This study describes the application of a genome scale transposon library combined with an experimentally validated genome-scale metabolic network of N. meningitidis to identify essential genes and provide novel insight to the pathogen's metabolism both in vitro and during infection.
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(2011) 'Chloroquine-enhanced gene delivery mediated by carbon nanotubes'. Elsevier Carbon, 49 (15), pp. 5348-5358.Full text is available at: http://epubs.surrey.ac.uk/7815/
Abstract
Polyethyleneimine-coated double-walled carbon nanotubes (DWCNTs) were used for dual gene and drug delivery, after loading the DWCNTs with the drug chloroquine, a lysosomotropic compound that is able to promote escape from the lysosomal compartment. Different forms of functionalization of the DWCNTs were examined in order to optimize this system. They included the testing of different treatments on DWCNTs to optimize the loading and delivery of chloroquine and the selection of a cationic polymer for coating the DWCNTs for optimum DNA binding and delivery. An acid oxidation treatment of DWCNTs was selected for optimum chloroquine loading together with polyethyleneimine as optimum cationic coating agent for plasmid DNA binding. Optimization of the conditions for choroquine-enhanced gene delivery were developed using luciferase expression as a model system. We have demonstrated that chloroquine-loading increases the ability of polyethyleneimine-coated DWCNTs to deliver functional nucleic acid to human cells. Cell viability tests have shown no cytotoxicity of the functionalized DWCNTs at the concentrations needed for optimum gene delivery. These results support the potential applications of this methodology in gene therapy.
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(2011) '¹³C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation.'. PLoS PLoS Pathog, United States: 7 (7)Full text is available at: http://epubs.surrey.ac.uk/184899/
Abstract
Mycobacterium tuberculosis requires the enzyme isocitrate lyase (ICL) for growth and virulence in vivo. The demonstration that M. tuberculosis also requires ICL for survival during nutrient starvation and has a role during steady state growth in a glycerol limited chemostat indicates a function for this enzyme which extends beyond fat metabolism. As isocitrate lyase is a potential drug target elucidating the role of this enzyme is of importance; however, the role of isocitrate lyase has never been investigated at the level of in vivo fluxes. Here we show that deletion of one of the two icl genes impairs the replication of Mycobacterium bovis BCG at slow growth rate in a carbon limited chemostat. In order to further understand the role of isocitrate lyase in the central metabolism of mycobacteria the effect of growth rate on the in vivo fluxes was studied for the first time using ¹³C-metabolic flux analysis (MFA). Tracer experiments were performed with steady state chemostat cultures of BCG or M. tuberculosis supplied with ¹³C labeled glycerol or sodium bicarbonate. Through measurements of the ¹³C isotopomer labeling patterns in protein-derived amino acids and enzymatic activity assays we have identified the activity of a novel pathway for pyruvate dissimilation. We named this the GAS pathway because it utilizes the Glyoxylate shunt and Anapleurotic reactions for oxidation of pyruvate, and Succinyl CoA synthetase for the generation of succinyl CoA combined with a very low flux through the succinate--oxaloacetate segment of the tricarboxylic acid cycle. We confirm that M. tuberculosis can fix carbon from CO₂ into biomass. As the human host is abundant in CO₂ this finding requires further investigation in vivo as CO₂ fixation may provide a point of vulnerability that could be targeted with novel drugs. This study also provides a platform for further studies into the metabolism of M. tuberculosis using ¹³C-MFA.
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(2011) 'Differential Producibility Analysis (DPA) of Transcriptomic Data with Metabolic Networks: Deconstructing the Metabolic Response of M. tuberculosis'. PUBLIC LIBRARY SCIENCE PLOS COMPUTATIONAL BIOLOGY, 7 (6) Article number ARTN e1002060 Full text is available at: http://epubs.surrey.ac.uk/184902/
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(2011) 'Optimising DNA binding to carbon nanotubes by non-covalent methods'. PERGAMON-ELSEVIER SCIENCE LTD CARBON, 49 (5), pp. 1775-1781.Full text is available at: http://epubs.surrey.ac.uk/7818/
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(2011) 'Acorn: a grid computing system for constraint based modeling and visualization of the genome scale metabolic reaction networks via a web interface.'. BMC Bioinformatics, England: 12Full text is available at: http://epubs.surrey.ac.uk/185976/
Abstract
Constraint-based approaches facilitate the prediction of cellular metabolic capabilities, based, in turn on predictions of the repertoire of enzymes encoded in the genome. Recently, genome annotations have been used to reconstruct genome scale metabolic reaction networks for numerous species, including Homo sapiens, which allow simulations that provide valuable insights into topics, including predictions of gene essentiality of pathogens, interpretation of genetic polymorphism in metabolic disease syndromes and suggestions for novel approaches to microbial metabolic engineering. These constraint-based simulations are being integrated with the functional genomics portals, an activity that requires efficient implementation of the constraint-based simulations in the web-based environment.
- . (2010) 'Carbon flux rerouting during Mycobacterium tuberculosis growth arrest'. WILEY-BLACKWELL MOLECULAR MICROBIOLOGY, 78 (5), pp. 1199-1215.
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(2010) 'Modeling and simulation of the main metabolism in Escherichia coli and its several single-gene knockout mutants with experimental verification'. BIOMED CENTRAL LTD MICROBIAL CELL FACTORIES, 9 Article number ARTN 88 Full text is available at: http://epubs.surrey.ac.uk/239125/
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(2010) 'Systems biology of the metabolism of Mycobacterium tuberculosis'. PORTLAND PRESS LTD Biochemical Society Transactions, 38 (5), pp. 1286-1289.doi: 10.1042/BST0381286Full text is available at: http://epubs.surrey.ac.uk/184901/
Abstract
Despite decades of research, many aspects of the biology of Mycobacterium tuberculosis remain unclear, and this is reflected in the antiquated tools available to treat and prevent tuberculosis and consequently this disease remains a serious public health problem. Important discoveries linking the metabolism of M. tuberculosis and pathogenesis has renewed interest in this area of research. Previous experimental studies were limited to the analysis of individual genes or enzymes, whereas recent advances in computational systems biology and high-throughput experimental technologies now allows metabolism to be studied on a genome scale. In the present article, we discuss the progress being made in applying system-level approaches to study the metabolism of this important pathogen.
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(2010) 'Physics and the mind Comment on 'Natural world physical-, brain operational-, and mind phenomenal-space-time' by Fingelkurts et al.'. ELSEVIER SCIENCE BV PHYSICS OF LIFE REVIEWS, 7 (2), pp. 250-251.Full text is available at: http://epubs.surrey.ac.uk/7370/
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(2010) 'Higher Dispersion Efficacy of Functionalized Carbon Nanotubes in Chemical and Biological Environments'. AMER CHEMICAL SOC ACS NANO, 4 (5), pp. 2615-2626.doi: 10.1021/nn100069kFull text is available at: http://epubs.surrey.ac.uk/735955/
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(2010) 'System-level strategies for studying the metabolism of Mycobacterium tuberculosis'. ROYAL SOC CHEMISTRY MOLECULAR BIOSYSTEMS, 6 (12), pp. 2363-2372.doi: 10.1039/c003757pFull text is available at: http://epubs.surrey.ac.uk/184934/
- . (2010) 'Uptake and Release of Double-Walled Carbon Nanotubes by Mammalian Cells'. WILEY-V C H VERLAG GMBH ADVANCED FUNCTIONAL MATERIALS, 20 (19), pp. 3272-3279.
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(2009) 'Identification of proteins from tuberculin purified protein derivative (PPD) by LC-MS/MS'. Elsevier TUBERCULOSIS, 89 (6), pp. 423-430.Full text is available at: http://epubs.surrey.ac.uk/7319/
Abstract
The tuberculin purified protein derivative (PPD) is a widely used diagnostic antigen for tuberculosis, however it is poorly defined. Most mycobacterial proteins are extensively denatured by the procedure employed in its preparation, which explains previous difficulties in identifying constituents from PPD to characterize their behaviour in B- and T-cell reactions. We here described a proteomics-based characterization of PPD from several different sources by LC-MS/MS, which combines the solute separation power of HPLC, with the detection power of a mass spectrometer. The technique is able to identify proteins from complex mixtures of peptide fragments. A total of 171 different proteins were identified among the four PPD samples (two bovine PPD and two avium PPD) from Brazil and UK. The majority of the proteins were cytoplasmic (77.9%) and involved in intermediary metabolism and respiration (24.25%) but there was a preponderance of proteins involved in lipid metabolism. We identified a group of 21 proteins that are present in both bovine PPD but were not detected in avium PPD preparation. In addition, four proteins found in bovine PPD are absent in Mycobacterium bovis BCG vaccine strain. This study provides a better understanding of the tuberculin PPD components leading to the identification of additional antigens useful as reagents for specific diagnosis of tuberculosis.
- . (2009) 'AFM imaging of functionalized carbon nanotubes on biological membranes'. IOP PUBLISHING LTD NANOTECHNOLOGY, 20 (43) Article number ARTN 434001
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(2009) 'Triple functionalisation of single-walled carbon nanotubes with doxorubicin, a monoclonal antibody, and a fluorescent marker for targeted cancer therapy'. PERGAMON-ELSEVIER SCIENCE LTD CARBON, 47 (9), pp. 2152-2160.Full text is available at: http://epubs.surrey.ac.uk/735957/
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(2009) 'The Genetic Requirements for Fast and Slow Growth in Mycobacteria'. PUBLIC LIBRARY SCIENCE PLOS ONE, 4 (4) Article number ARTN e5349 Full text is available at: http://epubs.surrey.ac.uk/184900/
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(2009) 'Towards the Immunoproteome of Neisseria meningitidis'. PUBLIC LIBRARY SCIENCE PLOS ONE, 4 (6) Article number ARTN e5940 Full text is available at: http://epubs.surrey.ac.uk/2561/
- . (2008) 'Mycobacterium tuberculosis strains disrupted in mce3 and mce4 operons are attenuated in mice'. SOC GENERAL MICROBIOLOGY JOURNAL OF MEDICAL MICROBIOLOGY, 57 (2), pp. 164-170.
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(2008) 'The acute-phase reactant C-reactive protein binds to phosphorylcholine-expressing Neisseria meningitidis and increases uptake by human phagocytes'. AMER SOC MICROBIOLOGY INFECTION AND IMMUNITY, 76 (3), pp. 1298-1304.doi: 10.1128/IAI.00741-07Full text is available at: http://epubs.surrey.ac.uk/351097/
- . (2007) 'Auxotrophic complementation as a selectable marker for stable expression of foreign antigens in Mycobacterium bovis BCG'. CHURCHILL LIVINGSTONE TUBERCULOSIS, 87 (6), pp. 474-480.
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(2007) 'Transcriptomic analysis identifies growth rate modulation as a component of the adaptation of mycobacteria to survival inside the macrophage'. AMER SOC MICROBIOLOGY JOURNAL OF BACTERIOLOGY, 189 (11), pp. 3969-3976.doi: 10.1128/JB.01787-06Full text is available at: http://epubs.surrey.ac.uk/184935/
- . (2007) 'Mycobacterium bovis BCG as a delivery system for the RAP-1 antigen from Babesia bovis'. ELSEVIER SCI LTD VACCINE, 25 (6), pp. 1104-1113.
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(2007) 'GSMN-TB: a web-based genome scale network model of Mycobacterium tuberculosis metabolism'. BIOMED CENTRAL LTD GENOME BIOLOGY, 8 (5) Article number ARTN r89 Full text is available at: http://epubs.surrey.ac.uk/184904/
- . (2006) 'Immunogenicity of Mycobacterium bovis BCG expressing Anaplasma marginale MSP1a antigen'. ELSEVIER SCI LTD VACCINE, 24 (37-39), pp. 6332-6339.
- . (2006) 'Generation of chemically unmodified pure single-walled carbon nanotubes by solubilizing with RNA and treatment with ribonuclease A'. WILEY-V C H VERLAG GMBH ADVANCED MATERIALS, 18 (12), pp. 1598-+.
- . (2006) 'Reduced toxicity of lipo-oligosaccharide from a phoP mutant of Neisseria meningitidis: an in vitro demonstration'. MANEY PUBLISHING JOURNAL OF ENDOTOXIN RESEARCH, 12 (1), pp. 39-46.
Conference papers
- . (2008) 'Dendritic cell responses to C-reactive protein-opsonised Neisseria meningitidis'. WILEY-BLACKWELL PUBLISHING, INC IMMUNOLOGY, Glasgow, SCOTLAND: Annual Congress of the British-Society-of-Immunology 125, pp. 124-124.
- . (2007) 'C-reactive protein binds to Neisseria meningitidis and affects macrophage responses to infection'. BLACKWELL PUBLISHING IMMUNOLOGY, Glasgow, SCOTLAND: Annual Congress of the British-Society-of-Immunology 120, pp. 20-20.
Mycobacterial research
The tubercle bacillus (Mycobacterium tuberculosis) infects approximately one quarter of the world's population and is responsible for three million deaths each year.
Mycobacterial research within the Microbial Sciences Group is focussed on understanding the pathogenic mechanisms that allow Mycobacterium tuberculosis to cause about three million deaths each year, and development of new vaccines to treat the disease.
We have recently initiated a new BBSRC-funded project to develop metabolic models of the TB bacillus and use these models to predict novel drug targets, particularly in persistent organisms. A web version of our model can be found here.
We currently have a Wellcome Trust-funded vacancy (to start autumn 2009) for a bioinformaticist to develop metabolic models of the TB bacillus.
Meningococcal research