Project Description

Synthetic Biology & Industrial Biotechnology offer a means of producing key organic molecules in a cleaner, greener and more sustainable way. In this research project, decarboxylase enzymes will be subjected to directed evolution (protein engineering/site-directed mutagenesis) in order to introduce new substrate and reaction specificity. The evolved enzymes will then be used either on their own, or as the initial catalyst in an artificial enzyme cascade to generate key high value intermediates for the pharmaceutical, chemical and food sectors. The effect of using a variety of non-aqueous reaction conditions on the evolved enzymes will also be examined. The PhD is one of a number funded by the Centre for Synthetic Biology at the University of Nottingham and you will join a cohort of students whose focus is on engineering proteins and metabolic pathways and will gain additional advanced training in synthetic biology. The work will be conducted in the Centre for Biomolecular Sciences which has state of the art facilities for protein manipulation and characterisation including X-ray crystallography, high field NMR, ITC, DSC, SPR and the School of Chemistry.

Project Description

The PhD project will be aimed at engineering Cuprividus necator to produce diols such as 1,4-butanediol and 1,3-propanediol from carbohydrate sources and then ultimately using carbon dioxide and hydrogen as sole carbon and energy source. One of the project objectives will be to design and implement synthetic metabolic pathways for the production of these diols. The project will aim to develop and apply combinatorial transcriptional engineering methodologies for synthetic metabolic pathway optimisation. The genetically encoded metabolic switch that enables dynamic regulation of pathway expression by adapting metabolic activity to the changing environment will be sought. Bacterial microcompartments, sort of organelles composed of a protein shell with several functionally interlinked enzymes compartmentalised on the inside, will be another alternative that will provide solutions for diol production improvement through the increase in local metabolite flux and reduced intermediate toxicity to the cell. Development and optimisation of the gas fermentation process for bioproduction will play an important part of the project.

Project Description

This project will integrate Life Cycle Assessment (LCA) and RRI methods to better understand the potential resource, environmental, and social implications of synthetic biology deployment. The integrated method will identify “hot spots” of high potential impact – e.g., feedstock requirements and procurement impacts, production impacts, product markets and social benefits and risks - and thereby inform ongoing technical research to reduce uneven or unintended consequences. The potential exists for social science to add value to LCA in a number of key ways, including accounting for social benefits and risks, incorporating diverse stakeholder perspectives and understanding ethical and value-driven drivers. The LCA-RRI framework will be applied to suitable case studies of relevance to the energy, chemicals, and/or pharmaceuticals sector.

Project Description

Zinc is an essential metal, required in ~30% of bacterial proteins, but is toxic at higher intracellular concentrations. Bacteria such as E. coli have evolved sophisticated zinc import and export systems controlled by transcription factors that repress the expression of genes encoding importer proteins (regulator Zur) or activate expression of zinc efflux (regulator ZntR). These regulators and the promoters they control represent a good example of fine tuning of cellular response to external zinc concentrations (1) and different Zur and ZntR regulated promoters have different affinities and transcription levels. We wish to study the levels of expression from engineered Zur and ZntR regulated promoters in response to zinc, so that a suite of promoters can be used to finely control gene expression in response to zinc levels in growth media. These promoters will be used to control gene expression in engineered bacteria using cheap zinc inducers and zinc chelators.

Project Description

The aim of this PhD will be to develop a new synthetic biology approach for the synthesis of biomimetic polymers, by genetic engineering of bacteria with iron reducing proteins to facilitate redox driven cell surface synthesis of polymers (FIG 1). The potential applications and therefore impact are broad in nature ranging from bacterial instructed routes in development of glycopolymer based drugs for treatment of disease, through to novel routes to manufacture biomimetic polymers to be used in tissue engineering and biodegradable plastics. Another interesting application is the ability to facilitate increases capture of cellular electricity in microbial fuel cells via directed growth of cell surface conducting polymers reducing our reliance on fossil fuels and petrochemicals.

Project Description

Hydrogen has real potential as a clean renewable fuel, producing water on combustion. Solar powered bio-hydrogen has several advantages; it is relatively harmless to the organisms producing it and is easily separated from the growth media. Photoautotrophic microbes like cyanobacteria can utilize cheap and plentiful sources of carbon and electrons for growth and hydrogen production making them self-sustaining production vehicles. However, diverting a high proportion of reducing power to hydrogen production poses significant challenges, exacerbated considerably by uncertainties in how the hydrogenase interacts with the electron transport chain. We have shown that the hydrogenase has two distinct physiologically-controlled localization mechanisms that partition it within the thylakoids, either dispersed uniformly through the thylakoids or aggregated into discrete puncta (Burroughts et al., 2014). Crucially, electron supply and hydrogen production depend on physiologically controlled localization. Determination of the molecular basis for control of hydrogenase location could thus pave the way to engineering improved cyanobacterial cells for solar-powered bio-hydrogen production and will form the basis of this project. Specifically the project will aim to determine the signal transduction pathway that controls Hox localisation in Synechocystis sp. PCC6803 and determine the influence of Hox localisation on electron transport pathways and hydrogen production. The student will utilise a wide range of techniques including knockouts, pulldowns and flash photolysis. There will also be the opportunity to evolve the enzyme to improve productivity through error prone PCR. The project will involve multidisciplinary collaborations across Kiel University (Dr Jens Appel), Warwick University (Prof Nigel Burroughs) and UCL (Prof Paul Dalby).

Project Description

Cupriavidus necator H16 is a Gram-negative, facultatively chemolithoautotrophic organism able to grow with organic substrates or CO2 / H2 under aerobic conditions. Its ability to grow on CO2 as sole carbon source makes it an ideal chassis organism for the sustainable production of platform chemicals from waste gases.

The aim of this project is to metabolically engineer Cupriavidus necator H16 for the sustainable production of value added chemicals from CO2 / H2. In particular, this project will evaluate producing chemical building blocks that are within three enzymatic steps from central metabolism, such building blocks having application as polymers or in sustainable agriculture.

Project Description

The project aims to create a rationally designed, multicellular biocatalytic structure, exemplified using a chiral amine biofactory. The multi-bacterial structure will mimic flow chemistry processes, and be capable of supplying the enzyme substrates, producing the biocatalyst and trapping the high-value product. This self-sufficient bio-factory will enable cost-effective and sustainable routes to important amine motifs, will represent an attractive approach for the industrial synthesis of chiral amines and provide proof of concept for the creation of other multicellular biofactories.

Project Description

The PhD project will use a synthetic biology approach to dissect the process of DNA replication in vivo and in vitro. DNA replication is the most fundamental task that proliferating cells must carry out, and begins at specific locations on the chromosome called origins. Archaea are the third domain of life, alongside eukaryotes and bacteria. Archaea are microbes renowned for living in extreme conditions such as acid pools and salt lakes. Haloferax volcanii comes from the Dead Sea and grows in saturated salt solutions. The enzymes that carry out DNA replication in archaea are strikingly similar to those used in eukaryotes. We have found that in the absence of origins, DNA replication depends instead on homologous recombination. The aim is to develop a simplified system for DNA replication based on homologous recombination. (1) To determine the minimum length for efficient replication by homologous recombination, we will construct large and small DNA molecules by genome engineering; (2) an inducible promoter will be used to regulate the expression of recombination genes, thereby facilitating the alternative mode of DNA replication; (3) a computational model will be developed to predict the rate of DNA replication, based on the length of recombining molecules and the rate of homologous recombination.

Project Description

This project investigates systems comprising of the halophilic protein alcohol dehydrogenase 2 (HvADH2) from H. volcanii and a broad range of ionic liquids (ILs) with diverse physicochemical properties. Addition of ILs offers intriguing prospects for stabilising globular proteins and their enzymatic function against deactivating reaction channels of folding intermediates and misfolds promoting irreversible protein aggregation. We examine the impact of highly heterogeneous molecular electronic structures of 13 ionic liquids on protein structure and bio-catalytic activity. Our pursuit is to trace back the role of specific functional groups and their influence on the microenvironment of HvADH2.

Project Description

Large-scale selective sequestration of specific bacterial species and/or strains from mixed populations is a long term goal in a number of academic and industrial sectors, spanning from biotechnology to healthcare. In recent years a range of technologies, spanning from microfluidics to hydrogels and lipid-silica structures have been developed to this aim, although they are generally limited to the confinement of small bacterial populations in defined locations. Within the context of synthetic biology, microbial engineering has been playing an increasingly important role in the biotechnological production of high-value chemicals, including pharmaceuticals.

This multidisciplinary project aims to develop a platform technology to selectively sequester specific microorganisms from complex mixtures once they are no longer needed for a given biosynthetic pathway, overcoming the problem of non-desirable reactions. This will involve (i) assembly and characterisation functional surfaces displaying appropriate ligands with properties which minimise unspecific bacterial binding, (ii) engineering of suitable bacterial strains with inducible displays of membrane receptors, and (iii) bacterial binding/sequestration from mixed populations to functional surfaces.

Project Description

The substrate and reaction scope of naturally occurring enzymes limits the production of high value, low molecular weight organic compounds by microorganisms. The availability of intracellular catalysts that facilitate non-natural transformations will therefore have enormous potential for the design of novel “intracellular retrosynthetic” pathways. To date, however, there are only few examples of chemical catalysts able to catalyse transformations in living organisms. To address this problem, we will design novel, non-natural imine reductases based on alcohol dehydrogenase scaffolds and evaluate their potential as building blocks for artificial metabolic pathways.

Project Description

The project aims to develop a platform for switchable gene expression control systems that will effectively allow either to fine tune gene expression in rationally designed metabolic pathways increasing efficiency of production in biotechnologically important microorganisms or to switch-off completely production of those enzymes, which are no longer required increasing metabolic flux towards desired chemical or other biotechnological product. The project will involve several multidisciplinary components: (i) engineering, for developing logic circuits and designing orthogonal logic gates; (ii) computational, for modelling and predicting behaviour of the system; (iii) molecular biology and genetic engineering, for engineering of gene expression mechanisms in metabolically versatile bacteria with high biotechnological potential such as Cupriavidus necator. The project will also train the PhD applicant in microbiology, analytical biochemistry and wide spectrum of synthetic biology approaches. 

Project Description

This synthetic biology project seeks to develop a principled approach to engineering microbes with minimal genomes adapted towards specified metabolic objectives. We will develop and combine novel wet lab (Vaud) and computational (Patel) techniques to create a high-throughput, broad host range pipeline capable of editing genes from target bacterial species to be transformed into designed chassis with the desired properties and enhanced exploitation potentials. Due to its relevance to industrial biotechnology, to its relatively close relatedness to Pseudomonas species for which we have extensive tools and expertise accumulated and to the innovative and challenging aspect of engineering it at the genomic level, we have selected Cupriavidus necator as the initial target organism.

Project Description

Methane is responsible for 10% of the world’s greenhouse gas emissions (on a CO2 equivalent basis), in part because it’s global-warming potential is twenty times greater than that of CO2 over a 100-year period. Using Synthetic Biology (SynBio) approaches, the studentship will be expected to design, implement artificial pathways and biological parts towards improving the performance of the recognised methanotrophic organisms (aerobic bacteria that utilise CH4 as the sole source of carbon and energy) that could potentially form the basis of a commercial process for the production of chemicals and fuels, particularly liquid transportation fuels.

Project Description

The principle aim of this PhD project will be the development of a suite of in-situ spectroscopic techniques that will enable the real time analysis and instrumentation of live fermentation cultures in laboratory and production scale gas fermenters.  We will probe bulk product concentration and develop novel techniques to allow the concentration of intra-cellular metabolites (both quantitative and qualitative) to be established while the fermenter is under growth conditions without the need for invasive and potentially problematic sample extraction.  The main analysis tools will include a comprehensive suite of vibrational spectroscopies (particularly infra-red (including NIR) and Raman techniques, both employing multiple detector fibre optic probes.  To complement in-situ spectroscopy we will also use in-situ chromatographic techniques to validate quantitative data.

Project Description

This synthetic biology project seeks to develop a principled approach to engineering microbes with minimal genomes adapted towards specified metabolic objectives. We will develop and combine novel wet lab (Vaud) and computational (Patel) techniques to create a high-throughput, broad host range pipeline capable of editing genes from target bacterial species to be transformed into designed chassis with the desired properties and enhanced exploitation potentials. Due to its relevance to industrial biotechnology, to its relatively close relatedness to Pseudomonas species for which we have extensive tools and expertise accumulated and to the innovative and challenging aspect of engineering it at the genomic level, we have selected Cupriavidus necator as the initial target organism.