Professor Martin Warren
Professor of Biochemistry/BBSRC Professorial Fellow
School of Biosciences
Professor Martin Warren joined the School of Biosciences in 2005. He is a member of the Industrial Biotechnology and Synthetic Biology Group and the Centre for Molecular Processing. Martin was born in Northern Ireland, brought up in County Down, and went to Portora Royal School, Enniskillen. He subsequently went to Southampton University where he read Biochemistry as an undergraduate (1981-1984). He stayed on in the Biochemistry Department to do a PhD with Professor Peter Shoolingin-Jordan, which initiated his interest in the genetics and biochemistry of tetrapyrrole biosynthesis.
After completing his PhD studies, he moved in 1989 to Texas A & M University, where he worked as a research associated with Professor Ian Scott FRS on vitamin B12 biosynthesis. In 1991 he took up a lecturing position in the School of Biological Sciences at Queen Mary, University of London, where he stayed until 1995 when he moved to a Senior Lecturer position at the Institute of Ophthalmology, Univeristy College London. He was promoted to Reader of Biochemistry in 1998 but then moved back to the School of Biological Sciences at Queen Mary in 1999 to take up a Personal Chair. In 2005 he moved to the University of Kent, where he is Professor of Biochemistry. In 2007 he was awarded a BBSRC Professorial Fellowship to work on the bioengineering of complex metabolic pathways.
He has published numerous articles on tetrapyrrole biosynthesis and the biochemistry underlying inherited retinopathies, as well as co-authoring a popular book on the link between tetrapyrrole biosynthesis and the madness of George III.back to top
Also view these in the Kent Academic Repository
Warren’s research envelops areas of chemistry, biology and history. His interests are firmly embedded in the biosynthesis and biology of the pigments of life, encompassing molecules such as heme, chlorophyll, vitamin B12, siroheme, coenzyme F430 and heme d1. With Dr Evelyne Deery he was one of the first to use synthetic biology as a means to probe biosynthetic pathways through the reconstruction of the whole cobalamin (vitamin B12) pathway in E. coli, an organism that does not possess the ability to make this molecule de novo. More recently he has elucidated a novel alternative heme biosynthesis pathway that is present in both archaea and sulphate reducing bacteria, where siroheme is hijacked as a substrate – a rare example of where one prosthetic group is cannibalised for the synthesis of another. His interest in vitamin B12 chemistry led to a study of how cobalamin is used for propanediol utilisation (pdu) in some bacteria, where remarkably the metabolic process is sequestered within a proteinaceous organelle called a bacterial micrcompartment, one of the largest protein-based complexes found in nature. Here, again, the entire pdu operon was cloned into E. coli and shown to orchestrate the construction of functional recombinant pdu microcompartments. Subsequently, empty organelles have been engineered through the targeting of new proteins and processes to these bodies. His interest in heme metabolism has also contributed to a new subject of scientific historiography. Here, he has studied George III, America’s last king, whose madness has been attributed to attacks of porphyria, an inborn error of heme metabolism. Through research in the Royal Archives in Windsor Castle and DNA analysis of some of the King’s descendants, coupled with a forensic analysis of the King’s hair, he has shown that it is likely that the much-maligned monarch suffered with variegate porphyria, worsened by the administration of antimony-based medication that was contaminated with arsenic.
BBSRC Funded research projects:
Bioengineering of complex metabolic pathways - Professorial fellowship 2008-2012
Vitamins are essential nutrients required by humans to complete their diet, and by definition are not made within their own body. Plants and vegetables are good sources of many vitamins. However, plants are missing one vitamin from their complement / vitamin B12. This is because plants neither make nor require vitamin B12 within their metabolism. In fact, vitamin B12 is unique among the vitamins in that it is the only vitamin whose synthesis is restricted solely to bacteria, as the ability to make this nutrient never successfully made the prokaryote to eukaryote transition. The consequence of this is that those on strictly vegetarian diets are prone to vitamin B12 deficiency - a state that is associated with a wide range of systems including megablastic anaemia, neurological disorders, and developmental problems in unborn babies. Vitamin B12 deficiency is also a problem in the elderly, where an increase in the level of B12 in the diet can alleviate the symptoms. There are thus strong medical reasons for increasing the levels of vitamin B12 in the diet.
In this research programme we wish to explore the limitations and consequences of engineering complex metabolic pathways into different organisms, taking advantage of the latest developments and technologies in metabolic engineering. We plan to take the genetic software that allows bacteria to make vitamin B12 and transfer it into bacteria that are unable to make B12, yeast and a higher plant, thereby conferring upon these organisms the ability to make this essential nutrient.
For bacteria, we wish to explore how the pathway can be enhanced for maximum vitamin production. Will increased levels of certain enzymes give increased metabolic flux, or will substrate bioavailabilty be limiting? These are questions that we will address in this application. For engineering into yeast, we have to look at a complex cloning procedure that ensures the genes have separate promoters and regulatory regions. For plants, we will take advantage of the fact that certain organelles within the plant, called plastids, have their own genetic material and in essence behave like bacteria within the plant. We will integrate the DNA from a bacterial species into the plastid and then monitor the level of the vitamin made during the growth of the plant. In fact, we will make a number of variants, of increasing complexity, to see how the plant is able to cope with this genetic modification.
This project is aimed at increasing our understanding of how biochemical pathways operate, how they are controlled and how they can be engineered to enhance the metabolic ability of the host cell. The results of the project will provide knowledge that can be used to develop new technologies and products for agriculture and bioremediation.
Unravelling the remarkable synthesis and mechanisms involved in the biogenesis of heme and heme d1 from siroheme
Co-applicants: Prof Stuart Ferguson (Oxford), Dr Mark Howard and Dr Stephen Rigby (Manchester) 2012-2014
This research aims to have a major impact on our understanding of the synthesis and evolution of a related family of heme-like molecules, and will result in a comprehensive addition to text book sections dealing with porphyrin metabolism. Hemes belong to a family of essential life pigment involved in many basic respiratory processes. This project aims to decipher a twenty-year mystery by unravelling the pathway leading to their synthesis. Specifically we aim to elucidate how heme and heme d1 are made in the Archaea and a number of eubacteria. Heme was previously thought to be synthesized along a single pathway from a compound called uroporphyrinogen III by a series of enzymes that modify the acidic side chains, oxidize the macrocycle and insert ferrous iron. However, we have recently demonstrated that heme and a structurally related macrocycle, heme d1, can be made by a completely novel pathway that involves the cannibalism of another modified tetrapyrrole, a compound called siroheme, the prosthetic group of sulphite and nitrite reductase.
There are many novel features to this project that make it compelling. The transformation of siroheme into heme and heme d1 requires a number of radical SAM enzymes coupled with a decarboxylase, involving some highly unusual steps that are unprecedented in biological chemistry. The aim of this research is to understand in molecular detail the step-by-step synthesis of hemes along this pathway.
Although the d1 heme is only found in the cytochrome cd1, the importance of this enzyme is becoming ever more evident as it vital for the environmentally important ANAMMOX process whereby ammonia is oxidised by nitrite under anaerobic conditions and has recently been implicated in playing a key role in the anaerobic oxidation of methane. It is believed that d1 heme has been adopted for respiratory nitrite reduction because it is tuned to permit nitrite reduction only as far as nitric oxide and not to allow the onward reduction to ammonia that is catalysed by siroheme. The discovery that siroheme is also a precursor for d1 synthesis therefore raises many interesting evolutionary and regulatory questions.
Overall, the outcome of this research will result in the composition of a molecular overture of events for a new heme synthesis route, explain more clearly the relationship between heme and siroheme and provide a neodarwinistic understanding of complex biosynthetic pathways. It will also help maintain the UK at the cutting edge of biomolecular science research and maintain the position our groups have as world leaders in this field.
Enzymes as traps in the elucidation of complex biochemical pathways
Co-applicants Prof Richard Pickersgill (Queen Mary), Prof Mike Geeves and Dr Mark Howard 2012-2015
In this research project we will apply a method that will allow a step-change in our ability to study complex biochemical pathways, provide molecular detail on fascinating enzyme mechanisms and to rewrite the metabolic control of pathways involving labile intermediates. The elucidation of biochemical pathways is a challenging area that is often complicated by low levels of inherently unstable metabolic intermediates. We have developed a method that allows for the isolation of enzyme-bound metabolites, permitting their characterisation and thereby providing an opportunity to gain atomic resolution of a number of fascinating enzyme-mediated transformations. The research is based on the finding that in some biochemical pathways the product of one reaction is passed directly onto the next in a process known as substrate channelling. Key to this is a tight association between an enzyme and its product, which allows for the isolation of highly stable enzyme-product complexes. We will exploit these properties to unravel the mysteries surrounding the biosynthesis of vitamin B12 (cobalamin). By using His-tagged enzymes of the pathway it is now possible to isolate many of the hitherto ephemeral intermediates, trapped and stabilised on the tagged enzymes as tightly bound enzyme-product complexes. Characterisation of these intermediates will allow the complete elucidation of the corrin pathway. Moreover, a combination of enzymology and X-ray crystallography will permit a detailed understanding of the mechanism of the enzymes that mediate the synthesis of the corrin framework, including the ring contraction process that involves the extrusion of an integral carbon atom in a reaction that has no parallel in nature. Our preliminary data is consistent with the B12 pathway operating by direct metabolite channelling. We outline experiments to investigate this further and to determine whether enzyme rather than substrate concentration controls this metabolic process.
Synthetic biology approaches to compartmentalisation in bacteria and the construction onovel bioreactors
Co-applicants Dr Dan Mulvihill, Prof Richard Pickersgill (Queen Mary), Dr Ian Blomfield 2010-2013
Synthetic biology has emerged from the development of techniques that allow for significant alterations in the metabolism/physiology of the cell to produce desirable products, including drugs, chemicals, vitamins, biofuels etc. Ultimately synthetic biology could lead to the construction of new metabolic pathways and even new life forms. A major advance in the area of synthetic biology would be the ability to make compartments in the cell to house specific metabolic processes. The development of such bespoke organelles would allow for greater control and regulation of the process, detaching the encased process from any negative influences within cellular metabolism. We have demonstrated that it is possible to synthesise such organelles in E. coli by the metabolic engineering of the 1,2-propanediol utilizing (pdu) operon, a complex that contains around 20,000 polypeptide subunits and has a molecular mass of approximately 300 mDa. More recently, we have shown that it is possible to make empty bacterial microcompartments (BMCs) by the coordinated overproduction of 5 gene products that compose the shell (outer casing) of the organelle. We have also demonstrated that proteins normally encased within the BMC are targeted to this empty vesicle and that other non-vesicle proteins can be incorporated to the BMC by fusing them onto target proteins. By labelling the proteins with GFP it has been possible to undertake live cell imaging of the organelles and evidence for movement between organelles via filaments has been observed.
We now wish to extend this study by directing specific enzymes to the BMC in order to make bespoke bioreactors. Such an approach will provide insights into the metabolic advantage of compartmentalisation. Engineering of the shell proteins will be undertaken to help in the rational design of a semi permeable shell with a broader substrate specificity. The intramolecular orientation of the shell proteins within the organelle will be investigated by a combination of antibodies and proteolytic processing. Detail on the intermolecular arrangement of shell proteins within the BMC will be obtained from studies using a range of different GFP-shell protein fusions and the analysis of the BMC by FRET. In this way we will be able to generate a model of the organelle and this, in combination with a number of other experimental approaches, will permit an investigation into how proteins are targeted and incorporated into the BMC. The rate of protein exchange in the organelle and the order of incorporation into the organelle will be defined not only using live cell imaging by also by employing FRAP. Detail on how the organelle is held within the cell will be investigated using a range of directed cell cytoskeleton mutants. The role of a Ras-type GTPase, PduV, in organelle dynamics will also be probed.
Mechanism of dimethylenzimidazole (DMB) synthesis and the metabolic engineering of a dietary useful form of cobalamin in Lactobacillus
Co-applicants Prof Gerald Richter (Cardiff), Prof Mike Geeves, Dr Gary Robinson 2010-2012
Vitamin B12, also known as cobalamin, is required for two essential enzymes in humans, methylmalonyl CoA mutase and methionine synthase and the lack of B12 prevents these enzymes functioning and results in pernicious anaemia. Although a form of the disease can occur in children, pernicious anaemia usually does not appear before the age of 30. The average age at diagnosis is 60. The disease occurs in all racial groups, but occurs most often in people of Scandinavian or Northern European descent. The disease may be manifest as a lack of vitamin B12 and / or an inability to absorb B12, both of which may be treated by vitamin B12 administration. Vitamin B12 supplements can be given but 1000 microg / day are required.
We ordinarily obtain vitamin B12 from our diet, particularly from meat and dairy products but not from fruit and vegetables, which do not contain this essential nutrient. Vitamin B12 deficiency results in a plethora of clinical manifestations including hematologic, neurologic and psychiatric disorders. Those people at risk of vitamin B12 deficiency include vegetarians –(where the absence of B12 in plants make vegetarians, especially vegans, susceptible to deficiency), the elderly (up to 30% of people over 70 have B12 deficiency and require supplements and, in some cases, intravenous injections of B12) and unborn babies (where B12 is a contributory risk factor in neural tube defects in unborn babies). There is thus a need to provide suitable foodstuffs with elevated cobalamin levels. Some countries are even contemplating adding B12 to flour in the same way that folate is already added.
Vitamin B12 is unique amongst the vitamins, being manufactured only by prokaryotes. It is produced commercially by fermentation using specific strains that have been selected for high B12 levels. Many strains and production methodologies have been used in the past but the principal methodology employed involves the co-feeding of a component of the vitamin called 5,6-dimethyl benzimidazole (DMB) to the culture medium. The synthesis and attachment of DMB is often rate-limiting in the biosynthesis of the pathway. Interestingly, as any trip to supermarket, health food shop or browse of the internet will demonstrate vitamin B12 can be purchased in a range of forms. However, many of these forms are useless as they do not contain the correct version of the vitamin, and include certain probiotic bacteria that are sold in liquid cultures and spirulina extracts, which are marketed as a source of vitamin B12 in the belief that, as cyanobacteria, they produce large amounts of the vitamin.
In these cases the proposed 'vitamin B12' is not cobalamin but is pseudocobalamin which, although structurally similar, is unable to be used by humans. The difference between the two compounds is the identity of the lower ligand with pseudocobalamin containing adenine and cobalamin containing 5,6-dimethylbenzamide (DMB). Expertise at Kent and elsewhere has contributed to our understanding of the synthetic pathway, leading to the characterisation of the protein responsible for the synthesis of DMB, enabling the production of true vitamin B12. The gene encoding this critical step (bluB) is present in certain bacterial strains but absent in others.
The overall strategy in the current research is to understand how DMB is made by the enzyme BluB. Subsequently, bluB will be transferred into a pseudocobalamin-producing probiotic strain that has GRAS (Generally Regarded As Safe) status. This is the first attempt to create a genetically engineered bacterium for use in probiotic formulations with the capability of providing sustainable vitamin B12 production in vivo.
back to top
- BI520 Metabolism and Metabolic Disease
- BI521 Metabolism and Metabolic Regulation
- BI629 Proteins