Understanding the role of sugar transporters in Mycobacterium tuberculosis

Tuberculosis (TB) is caused by the bacterium Mycobacterium tuberculosis (Mtb). TB is one of the leading causes of death worldwide from a single infectious agent. In 2011, 1.4 million people died from TB and 8.7 million people became infected. The problem of TB is not confined to developing countries and in the last decade cases of TB have doubled in the UK with London now known as the TB capital of Western Europe.

Treatment of TB requires a lengthy, complicated drug regimen, which means that many patients fail to follow the recommend course of treatment. Failure to comply with the drug regimen, in turn, has led to the evolution of multi-drug resistant and extensively drug-resistant TB (MDR-TB and XDR-TB). MDR- and XDR-TB strains are difficult and costly to treat with existing antibiotics and in some instances, few, if any, therapeutic agents remain. In conjunction with HIV infection, this deadly infection now presents us with a global time bomb that could devastate societies across the globe.

M. tuberculosis is unique from most bacteria that cause infection in that it has a distinctive, unusual cell wall. The cell wall has an unusually high fat and sugar content that provides a barrier and protects Mtb from the immune defence system. The cell wall prevents many common antibiotics, such as penicillin, from being effective. As a result of this protection the bacteria are able to survive and persist undetected in the immune system for decades. The disease can 'awake' when the immune system weakens, for example in old age or HIV-infection.

The way that Mtb causes infection and the energy sources it uses whilst surviving within human cells and tissues are still poorly understood and it is thought that Mtb survives on a diet of fats and sugars. From scrutiny of its genome, it appears that Mtb has five potential systems that can be used to transport sugars into the cell; however, currently, we do not know the sugars that these systems select and uptake. Studies suggest that some of these transporter systems are essential during the early stages of infection and therefore could potentially be used as drug targets.

Mtb has fewer sugar transporters than other bacteria, suggesting that Mtb has evolved to survive within niche environments within the human body where a limited selection of sugars is available for it to use as energy sources. Understanding how Mtb selects and uses or recycles the sugars will lead to new insights into the survival mechanisms of this deadly pathogen. Fully understanding the role of these transporters is fundamental to enable us to develop new drugs to treat this disease.

The work will be carried out at the University of Warwick where a two-pronged approach to the research will be undertaken. Firstly, a detailed biochemical study of the proteins involved in sugar recognition will be performed. Secondly, genetic studies will be used to selectively remove the function of each of the potential sugar transporters to test if the Mtb mutant strains are still able to grow, uptake sugars and cause infection. Fully understanding the role of each of the sugar transporter systems in Mtb will deliver an improved understanding of the energy sources that are available to the bacterium during infection and factors that enable Mtb to survive within humans.

This research has important medical implications and has significant potential to impact on human health. During these studies there is the potential to look for compounds that inhibit the uptake of sugars by Mtb, which in turn could lead to novel, potent anti-tubercular agents that are urgently needed to improve the outcome of patients with TB.

The aim of the project is to elucidate the role of the putative sugar transporters in Mycobacterium tuberculosis (Mtb). These transporters have been shown to be important in the intracellular survival of Mtb. In order to fully determine the role of these systems a two-pronged approach will be undertaken:

1) Clone, express and purify enzymes involved in sugar uptake in Mtb, with an emphasis on biochemical characterisation, structural biology and drug development
2) To assign physiological functions to genes involve in sugar transport by generating null and conditional mutants in Mtb.

Recombinant proteins will be produced for the proteins involved in sugar recognition and the substrate specificities and binding constant determined (ITC, NMR, DSF). I will determine the crystal structures, with and without ligands, of these proteins and screen for small molecule inhibitors.

In parallel, I will generate null and conditional mutants of the genes involved in sugar transport. The growth kinetics of the mutants monitored, the effects of sugar uptake determined and the accumulation and incorporation into cell wall components of Mtb of sugar metabolites assessed using radiolabelled sugars. Mutant strains will be examined both in macrophages and in mice in order to assess changes in virulence and to determine the functional role of the sugar transporters in Mtb infection.

Full biochemical and genetic characterisation of these transport systems will be a major advance in understanding Mtb infection and will lead to the development of inhibitors.

The research has significant medical implications with the potential to determine the role of these systems in virulence of Mtb and to validate targets for TB therapy, leading to new anti-tubercular agents. The increasingly prevalence of drug resistant TB means the identification of novel treatment targets is of vital importance to address this global health issue.

Throughout the project the potential for exploitation and dissemination of the research will be carefully monitored.

Tuberculosis (TB) is a significant global health problem and the increasing incidence of antibiotic resistance is an issue that it is vitally important to address.

This research will address the current gap in our understanding of nutrient acquisition in Mtb. In particular, the proposed work will lead to an increased knowledge as to the physiological substrates of the sugar transporters in Mtb and will provide greater understanding of the mechanisms of uptake of sugars, virulence factors and mechanisms for the intracellular survival and establishment of infection of Mtb. The biosynthetic pathways to be investigated in this research proposal have the potential to be important novel targets for anti-tubercular therapy will therefore generate significant medical outcomes on a global scale. I will work closely with Warwick Ventures, the University of Warwick technology transfer office, to ensure that any intellectual property relating to potential targets and compounds that have potent anti-tubercular properties will be suitably protected. In addition I will also work with Warwick Ventures to ensure that such compounds and targets can be commercially exploited and translated into effective treatments in the longer-term.

Greater understanding of Mtb infection and potent anti-tubercular agents will primarily improve the health of nations in the developing world, along with immigrant populations in the UK. More people will be able to work if they are not suffering from TB infection and along with improving qualify of life, this will create wealth and economic prosperity for developing nations. TB is among the three greatest causes of death among women aged 15-44 and preventing mothers from dying will improve the health and welfare of subsequent generations. Furthermore, in many countries TB is a disease with social stigma attached and improving public awareness of TB, coupled with potential therapeutic agents will be beneficial to easing these cultural boundaries.

The research will also seek to improve public awareness and understanding of TB. Press releases through the media to the general public will be promoted and made available when necessary. TB is predominantly a problem in large cities in the UK and therefore educating the general public and schools through outreach programs at the University of Warwick, in which the research will be promoted, will be beneficial and undertaken during the time-frame of the award.

Involvement in the Tuberculosis Drug Discovery Consortium (TBD-UK) will enable outcomes of the research to be disseminated to policy makers such as the All Party Parliamentary Group on Global Tuberculosis and non-profit organizations such as the TB Alliance and Stop-TB partnership, a World Health Organisation initiative to eliminate TB as a public health concern.

The range of approaches will provide the research technician and fellow with a range of professional skills suited to career development in both academic and industrial settings, contributing to skilled workers in the UK work force. The research will also be of international standing and along with the recent investment in Infection and Immunity at the University, help to further establish the University of Warwick as a leading centre for infectious disease research in the UK that will be highly competitive in attracting excellent researchers.