The goal of this project is to understand the protein-protein interactions involved in the assembly of the bacterial type III secretion needle apparatus. This apparatus resembles a syringe on the bacterial surface and is used by many pathogens to inject virulence factors into target cells to initiate human diseases. The needle-like assembly consists of about 120 copies of identical proteins that are arranged in a superhelical manner, and the apex of this needle is capped by tip proteins. Over the past two years, we have completed the NMR structures of two needle monomers: BsaL from Burkholderia pseudomallei, a pathogen associated with biowarfare, and PrgI from Salmonella typhimurium, a pathogen associated with food poisoning. Our aim is to elucidate how the needle apparatus is assembled by determining the precise protein-protein contacts involving the needle and tip proteins. We will use NMR with electron microscopy and molecular modeling to determine the structure of the assembled needle. The aims of this project are: determine how the needle is assembled, determine how the tip protein docks on the needle, and use mutagenesis and microbiological assays to correlate the structural results to needle assembly and virulence in vivo.

Another project is to determine the protein-protein and protein-RNA interactions involved in the assembly of hantavirus. Hantaviruses infect tens of thousands of people worldwide. Rodents are the primary reservoir and humans are infected by inhalation of dust or aerosol contaminated with the excreta of infected rodents. The lethality, worldwide distribution, and aerosol transmission make the hantaviruses potential threat to public health and safety. There are no vaccines nor specific antiviral therapies approved for the hantaviruses, thus, there is a critical need to develop specific antiviral therapies. The RNA genome of hantaviruses encodes four proteins: nucleocapsid, RNA polymerase, and two membrane glycoproteins, G1 and G2. G1 has a glycosylated ectodomain and a 135-residue cytoplasmic tail. The G1 tail is important in viral assembly and the evasion of host response against viral infection; it binds the viral ribonucleoprotein as well as cellular proteins. Our goal is to determine the protein-RNA and protein-protein interactions of the G1 tail with respect to viral assembly and host-pathogen interaction. This knowledge is important not only in understanding how the virus is assembled, but also in developing specific antiviral agents.