Projects / Programmes
Cylcic nucleotide phosphodiesterase from Mycobaterium tuberculosis: structure and function
| Code |
Science |
Field |
Subfield |
| 1.05.00 |
Natural sciences and mathematics |
Biochemistry and molecular biology |
|
| Code |
Science |
Field |
| B230 |
Biomedical sciences |
Microbiology, bacteriology, virology, mycology |
tuberculosis, cylcic nucleotides, phosphodiesterases, cell signaling, crystal structure
Researchers (10)
Organisations (2)
Abstract
One third of the world’s population is infected by the causative agent of tuberculosis, Mycobacterium tuberculosis (Mtb), resulting in close to 2 million deaths a year. As a pathogen, Mtb has developed sophisticated mechanisms to evade the host immune system. These could involve the use of cyclic nucleotide (cAMP/cGMP =cNMP) dependent signaling systems, since the Mtb genome encodes a large number of functional nucleotide cyclases, the enzymes that synthesize cNMPs.
Using bioinformatics, biochemical and structural approaches we have recently, together with our collaborators from Inida, identified and characterized the gene Rv0805 in the genome of Mtb coding for the only cNMP-phosphodiesterase so far found in this mycobacterium. The Rv0805 protein is a class III cNMP- phosphodiesterase, hydrolyzing cyclic NMPs to respective nucleotide monophosphates (NMPs) in vitro and in vivo. It is a dimeric protein and the crystal structure of the catalytic core reveals typical calcineurin-like metallo-phosphoesterase fold. The active site of Rv0805 contains two catalytically and structurally important metal ions, Fe3+ and Mn2+.
With this application we are proposing the extension of our studies in order to: (a) Determine the crystal structure of the full length Rv0805; (b) Study the catalytic promiscuity of Rv0805; (c) Study the interactions between Rv0805 and other bacterial proteins; (d) Identify and study Rv0805-related proteins in other bacterial and eukaryotic organisms.
This project has been designed in order to obtain knowledge on the role of cNMPs and the enzymes involved in their metabolism, in physiology and pathogenesis of mycobacteria. Our research partner in this project in one of the leading Slovenian pharmaceutical companies, Lek Pharmaceuticals d.d. We believe that the complementary expertise of the groups collaborating on this project can be of a great advantage not only for a successful development of the project, but also for the potential medicinal and pharmaceutical application of results.
Significance for science
Transduction of signals mediated by cyclic nucleotides is relatively poorly explored in mycobacteria. Since there is growing evidence that cyclic nucleotides may be involved in mycobacterial pathogenesis, this interesting scientific niche calls for more research projects. Seven years ago we started collaboration on this topic with a group of Prof. Sandhya S. Visweswariah from Indian Institute of Science, Bangalore, India, who has been successfully working in this field for several years. Together we identified to date the only known cyclic nucleotide phosphodiesterase of Class III, Rv0805 from Mycobacterium tuberculosis. Results coming from this work represent the first detailed analysis of the member of this class of enzymes. The main goal of our work is to contribute to understanding of the action of these enzymes in mycobacteria as well as in the cross talk with their eukaryotic hosts. Results comimg from this project, represent steps closer to these objectives. Particularly interesting are those showing how small changes in amino acid sequences between the enzymes of the same superfamily or class but from different organisms affect their substrate specificities. Furthermore, our data that show localization of Rv0805 enzyme at the bacterial cell wall and how Rv0805 affects also the cell wall permeability. It is well known that a unique mycobacterilal cell wall is a very important weapon in attacking the host and is thus important for bacterial survival in different environments. Therefore the cell wall- attached molecules are potential drug targets. This is an important basis for our future studies which will include further comparison of evolutionary different organisms and the role of the proteins from Class III in them. In fact, our recent results on Rv0805 ortholog from mammals indicate how the nature used a particular fold of the polypeptide chain and highly conserved active site residues, and modified them to the extent that enables this molecule a different biological role. Furthermore, the mammalian ortholog of Rv0805 is also very interesting from scientific as well as medical point of view, since there is growing evidence that deletion of this gene in humans may be involved in syndromes like mental retardation which can be observed in some newborn children (the WAGR syndrome).
Significance for the country
1) New scientific achievements, originating from our recent work, contribute to better understanding of the basic biological processes in (pathogenic) mycobacteria, and in the same time help to understand similarities or differences in biological processes of evolutionary different organisms, many of which are hosts for these bacteria. Understanding of the basic mechanisms of the biological processes in mycobacteria as well as of the relationship pathogen-host, is very important also from the aspect of medicine and pharmaceutical companies (development of drugs), especially in the case of the diseases such as tuberculosis. Namely, the causative agent of this disease, Mycobacterium tuberculosis, has evolved strains which are extremely resistant towards existing antibiotics, and has been therefore becoming again one of the major death threats (based on the disease) in the world. This problem has been historically acute for the countries of the World's South and East, with poverty being one of the main reasons. However, with the major world economy crisis and increasing social differences leading into increased population of poor people, wars and refugees, deadly diseases are also spreading back to the developed countries. Therefore prevention as well as appropriate curative applying also scientific discoveries is important in Slovenia as well. As already mentioned above, we also hope that our discoveries on mammalian ortholog of Rv0805 will help understand the complexity of the WAGR syndrome, which affect fetus and later on newborns, with a deletion in the gene encoding this protein. With our studies we primarily wish to learn how this protein works under normal conditions, and therefore what would biochemically as well as medically mean if this protein was absent in the organism.
2) Promotion of Slovenia and its science: scientific meetings (active contributions in forms of invited lectures and/or posters), publications in the international scientific journals and collaboration with research groups around the world. Collaboration on this project with the group of Prof. Sandhya S. Visweswariah from Indian Institute of Science, Bangalore, India, who visits our group regularly once a year, as well as other members of her research group. This collaboration is also supported by a bilateral grant.
3) Contribution to Slovenian education system and availability of new job positions: Dr. Marjetka Podobnik, the leader of this project, is a mentor to PhD and diploma students. One of her PhD students, Urška Dermol, has been working on the topics of this project. Dr. Podobnik was also a mentor to Petra Draškovič, who finished her PhD in March 2009. Her research was oriented towards inositol pyrophosphates, another group of potential second messengers. Since May 2009, Dr. Podobnik is also a supervisor of a postdoctoral associate, Dr. Katja Rebolj.
Most important scientific results
Annual report
2008,
2009,
final report,
complete report on dLib.si
Most important socioeconomically and culturally relevant results
Annual report
2008,
2009,
final report,
complete report on dLib.si
