Molecular Chaperones and Immunity
Design of Recombinant Protein-Peptide Immunomodulators
Tech Area / Field
- BIO-CGM/Cytology, Genetics and Molecular Biology/Biotechnology
- MED-DRG/Drug Discovery/Medicine
3 Approved without Funding
Institute of Immunological Engineering, Russia, Moscow reg., Lyubuchany
- Russian Research Plague Institute "Microbe", Russia, Saratov reg., Saratov
- University of Turku / Finnish-Russian Joint Biotechnology Laboratory, Finland, Turku\nLabmaster Ltd., Finland, Turku\nMichigan State University, USA, MI, East Lansing\nUniversity of Reading, UK, Reading
Project summaryStudying the molecular mechanisms of the virulence of pathogenic micro-organisms, particularly those, which are responsible for their ability to persist in the organism of the immune host, is one of the most urgent problems in biology and medicine. It is necessary to solve this problem in order to develop effective means for prevention and treatment of infectious diseases.
Lately cardinal changes have taken place in understanding the molecular mechanisms of virulence of Gram-negative bacterial pathogens. These changes are caused, first of all, by the discovery of the type III secretion system, providing direct communication between the bacterium cytoplasm and the host target-cell cytoplasm when they are in tight contact. With the assistance of this system bacteria secrete effector proteins, disturbing normal functioning of the target-cell, into the host cell.
To realise the function of the type III secretion system, the pathogen and the host cell must have a close contact which is obtained through specialised surface structures (pili, fimbria, protein capsules and others). At present a considerable progress has been made in studying the structure and the function of bacterial operons, responsible for forming the above-mentioned surface structures, which are often associated with virulence.
The most common property of the operons, responsible for the formation of virulence-associated surface structures of Gram-negative pathogens, is that the genes encoding periplasmic molecular chaperones and outer membrane usher-proteins are always present in them. The main functions of the first group of the proteins are their assistance to correct folding of surface structure subunits, prevention from non-productive association, transmission of the subunits to the usher-protein for their further translocation on the bacterium surface.
Previously the authors of the Project have cloned and sequenced genes of caf1 operon, responsible for forming a capsule on the surface of Y. pestis. Caf1 operon consists of caf1, caf1M, caf1A and caf1R operons, encoding capsular antigen F1 (Caf1), periplasmic molecular chaperone Caf1M, molecular usher protein Caf1A and Caf1R (a protein-regulator of caf1 operon expression), respectively.
The principle protein of the Caf1 capsule is one of the pathogenicity factors and the main Y. pestis immunogen. Recently we have obtained some preliminary data on the interaction of the dimeric form of Caf1 protein with one of the key cytokines in the immune system - huIL-1. At the same time it has been established that the polymerous form of Caf1 does not interact with huIL - b1. Possibly, the interaction of Caf1 dimer with huIL - b1 plays an important role in the plague pathogenesis at the early stages of capsule formation in bacteria when infecting the host. At present there are some reports indicating existence of Y. pestis virulent strains with a serologically atypical capsule (F- phenotype). It is supposed that the atypical capsule is formed in bacteria if there are defects at the level of caf1M gene. To develop a vaccine effective against F- strains of Y. pestis it is necessary to elucidate the nature and molecular mechanisms of forming the serologically atypical capsule, formed by Y. pestis strains defective in Caf1M gene. To search for new pharmaceutical preparations for treatment of plague, it is necessary to study the molecular mechanisms of the interaction of Caf1 protein with cytokines of the IL-1 family.
The objective of the studies under the Project is to study the chaperone-dependent mechanism of antigenic changeability of plague agent Yersinia pestis as a way to improve the molecular antiplague vaccine.
The scientific novelty of the proposed studies is in elucidating the peculiarities of the molecular functioning mechanisms of operons of Gram-negative pathogens (using caf and psa operons of Yersinia pestis as an example) responsible for the formation of virulence-associated surface structures with atypical properties and influencing the chaperone-dependent mechanism of the antigenic changeability of these pathogens. In particular, some data would be obtained on the interaction of the products of caf and psa expression. We also plan to study the molecular mechanisms of the interaction of different forms of capsular Caf1 with cytokines of the IL-1 family.
To achieve the objectives set in the Project we are going to use a complex of modern research methods, including gene engineering methods, site-directed mutagenesis, NMR-spectrometry, electron microscopy, differential adiabatic scanning microcalorimetry, circular dichroism, spectrofluorimetry, molecular computer modelling, radioreceptor analysis. The biological activity of the mentioned compounds will be studied using a complex of biochemical, immunological and pharmacological methods.
The Project execution would make a significant contribution to the development of fundamental and applied biology and medicine as well as to the solution of a social problem, - prevention of bioterrorism, a serious threat for the world. (Glenn E. Schweitzer, et. al., 1998; Richard A. Falkenrath et. al., 1998). Besides, the Project execution would give the scientists and engineers from the Institute of Immunological Engineering, Joint-Stock Company “Biopreparat”, who fulfilled earlier state orders of the Ministry of Defence, an opportunity to apply the accumulated experience and the international relations, they obtained executing Project 091-94, to carrying out fundamental and applied studies, connected with the solution of international research and technical problems in the field of biology and medicine for peaceful purposes, to creating long-term perspectives of fruitful activity within the international scientific community.