It is proposed to determine the three-dimensional structure of pertussis toxin by high resolution x-ray crystallography. Pt is a protein exotoxin produced by Bordetella pertussis, the bacterium that causes the infectious disease known as whooping cough, a disease affecting over 60 million infants and leading to over one million deaths per year world-wide. A specific objective of this research is to define those structural features of PT that are important in eliciting the protective immune response after natural infection or vaccination. Although protective vaccines prepared rom unfractionated B. pertussis cells have long been available, they are often associated with adverse side effects including rare cases of neurological impairment. The development of improved vaccines, free of deleterious complications, requires the definition of the minimal molecular structures against which the protective response is mobilized. Since pertussis toxin is considered a major virulence factor in the disease, the specific immune response to PT is almost certainly essential for protection. A high resolution structure of Pt in conjunction with available genetic and biochemical data would allow recognition of structural features involved in the immunogenicity, enabling a distinction to be made between those regions of the molecule conferring protective immunity from those simply eliciting a response. Another important reason for obtaining more detailed structural information concerning PT is that its mode of action is the interference with a critical component of the fundamental signal transduction machinery of eukaryotic cells. The current unifying hypothesis is that a wide variety of receptor mediated events at the membranes are effected by a class of proteins known as the G proteins. It is known that Pt catalyses the covalent modification of G proteins by transferring ADP-ribose from NAD+. Finally, an important aim of the work is to better understand the basic principles involved in mechanisms by which proteins such as the A subunit of PT are internalized by cells at the membrane after interaction with translocating complexes like the B subunit of PT. A three-dimensional structure of PT could lead to more effective and safest vaccines against whooping cough, provide a basis for designing cytotoxic agents coupled to cancer cells targeting antibodies, and contribute to our improved understanding of basic receptor biology and oncogenesis.
