PCHPI Project Summary Essentially every step in the HIV life cycle interfaces intimately with the host cell machinery. The Pittsburgh Center of HIV Protein Interactions will focus on the steps and interactions that occur with the host after engagement of cell surface receptors and membrane fusion through integration of the viral genome into that of the host, the so called “early events”. Several essential molecular interactions and enzymatic activities occur within this time window, necessary for productive progression of the viral life cycle. Thus, it represents a pivotal period in the infection process, during which the susceptibility of the virus to disruptive interventions is likely to be high and little explored. Broadly speaking, the processes that we will focus on include uncoating (the process by which the capsid disassembles), reverse transcription, evasion from innate immune factors, nuclear entry and integration. Given the importance of the capsid structure and its interactions for many of these processes, we will also explore maturation and in particular formation of the capsid core. We plan to build on our successes and apply the extensive and complementary experimental expertise of our team to carry out 1) biochemical and high-resolution structure studies of individual proteins and complexes, 2) biochemical, biophysical, and proteomics analyses to identify novel interactions and complexes, 3) virology and imaging studies to understand protein function in the context of the cell and virus infection, and 4) computational analyses to elucidate the physical basis of capsid formation and capsid interactions with binding partners. Broadly, the program comprises projects on capsid interactions, the engagement of Vpr with the DNA repair machinery, and retroviral intasome structure. Further, we plan two new Technology Development Programs, one to develop nanocrystal screening specifically for MicroED and the other to develop ultra-fast MAS NMR methods for structure determination of native particles.