Time-resolved studies of HIV-1 capsid protein self-assembly

National Institute Of Diabetes And Digestive And Kidney Diseases

Investigators

Abstract

Funds for this new project were received from the Office of AIDS Research in April 2024, through the OAR Innovation Program. Funds from OAR were used to order a microwave source (high-power extended interaction oscillator), which is required for the planned solid state NMR measurements on frozen solutions of CA self-assembly intermediates. Additional funds from the NIDDK Intramural Research Program were used to order stopped-flow mixing equipment, which is required for time-resolved light scattering measurements. These measurements will allow us to identify optimal conditions (e.g., CA and IP6 concentrations and self-assembly times) for the formation of oligomeric intermediates. In FY2025, the stopped-flow mixing equipment was received, installed and tested. HIV-1 CA samples were prepared with uniform 15N and 13C labeling of the entire protein sequence, with uniform labeling of only the C-terminal domain, and with 15N and 13C labeling only of Met residues in the N-terminal domain and only of Leu and Thr residues in the C-terminal domain. Split intein methods were used to prepare the latter two samples. The high-power microwave source has not yet been received from the manufacturer, but is expected to arrive early in FY2026. At that point, experiments to identify and characterize intermediates in the HIV-1 CA self-assembly process will begin in earnest.