Molecular analysis of complement activation via the classical pathway

The human immune system uses two main weapons to counter invading pathogens: adaptive immunity, which includes the production of antibodies that develop over time to target invading bacteria; and innate immunity, which is ever present and recognizes conserved parts of bacteria, such as their cell walls. The complement cascade provides a bridge between innate and adaptive immunity: it destroys bacteria directly via innate processes and helps to direct and stimulate an effective adaptive response. Because of its central role, complement dysfunction is associated with a range of human diseases, including severe bacterial infections, kidney disease and autoimmune diseases such as lupus and rheumatoid arthritis. One of the main pathways of complement activation is known as the classical pathway, in which complement proteins bind to bacteria and set off a chain of reactions directed at killing them. In this grant proposal, we aim to develop a thorough understanding this pathway by studying the way in which the components interact with each other to initiate the reaction cascade. We have already established methods for producing key elements using recombinant DNA technology so that their mode of action can be investigated in a systematic way. As well as providing novel information that will help us to understand how complement works, this project will also foster the development of novel reagents which can be used to change complement activity. For example, inhibitors of complement could be used to prevent tissue damage in conditions where complement activates at the wrong time or place. Similarly, modified components can be used to bind and neutralize specific targets such as antibiotic resistant bacteria or diseased cells.

The classical pathway of complement activation provides a frontline defence against invading pathogens. It is a central effector of natural protection, one of two current research priorities of the MRC. It neutralises invading cells directly via antibody-dependent and -independent mechanism and elicits a range of cellular responses designed to protect the host and eliminate foreign material, including inflammation, phagocytosis and stimulation of an effective adaptive immune response. Despite its important roles, the mechanisms underlying the activation and regulation of this branch of the immune system are relatively poorly understood. The goal of the proposed work is to understand how molecular interactions control complement activation via C1, the initiating complex of the classical pathway. Biochemical and structural approaches are specifically designed to identify detailed binding and structural information. The results of this study are expected to be of significance to both basic and applied applications. Understanding the fundamental issues of recognition in complement activation has general practical applications in protein-protein interactions and protein engineering and lays the foundations for the development of therapeutic agents designed to prevent complement-mediated host damage for use in the treatment of inflammatory diseases, such as rheumatoid arthritis and in ischemia-reperfusion disorders. As part of this project, short constrained peptide mimics will be developed, synthesized and characterized. These will serve as specific inhibitors of C1q-C1rs interactions, providing a novel set of reagents to dissect the complex biological processes that these proteins mediate. We also plan to use our knowledge of complement interactions to engineer novel specificities into the framework of C1qs, MBLs and ficolins, thereby altering target specificity whilst retaining the ability to activate complement and stimulate the host?s immune system. By targeting bacterial cells and endogenous epitopes associated with disease, these immunologically active reagents will offer a novel approach to tackle disease.