Poster abstracts

Poster number 12 submitted by Varun Dewan

Discovery of Cyclic Peptide Inhibitors Against HIV-1 Capsid

Varun Dewan (Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210), Tao Liu (Department of Chemistry, The Ohio State University, Columbus, OH 43210), Kuan-Ming Chen (Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota 55455), Hiroshi Matsuo (Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota 55455), Dehua Pei (Ohio State Biochemistry Program, Departments of Chemistry and Biochemistry, and Center for Retroviral Research, The Ohio State University, Columbus, OH 43210), Karin Musier-Forsyth (Ohio State Biochemistry Program, Departments of Chemistry and Biochemistry, and Center for Retroviral Research, The Ohio State University, Columbus, OH 43210)

Abstract:
The Capsid (CA) domain of HIV-1 Gag polyprotein plays a critical role in both the early and late phase of viral replication. In the early phase of HIV-1 infection the CA coat disassembles and releases the RNA genome into the host cell where it is reverse transcribed and integrated into the host genome. Upon translation, the CA domain of the Gag polyprotein binds to human Lysyl-tRNA synthetase (LysRS) to form a tRNALys primer packaging complex. Further, as the virus buds from the host cell membrane during the late phase, the proteolytic cleavage of the Gag protein leads to the formation of the characteristic conical core formed by closed hexameric array of the viral CA protein in the mature virus. Being versatile in nature makes CA an important antiviral target for disrupting the assembly of HIV-1. Small-molecule inhibitors against HIV-1 CA, commonly referred to as capsid assembly inhibitors (CAI), have potential clinical applications and hence may serve as useful molecular probes in biomedical research. In this work, a robust method has been employed for the high-throughput synthesis, screening and identification of cyclic peptidyl ligands against macromolecular targets. Support-bound cyclic peptide libraries containing randomized amino acid sequences and different ring sizes (theoretical diversity of 1.2 × 107) were synthesized and screened against CA and the monomeric form of the CA-terminal domain (WM-CA CTD). Cyclic Peptide 2 and 4 (CP2, CP4) out of six selected cyclic peptides showed strong binding (KD ~ 500 nM) to both CA and WM-CA CTD in vitro. The scrambled variants of CP 2 and CP 4 resulted in no binding to either CA or WM CA-CTD suggesting a sequence specific mode of interaction. CP2 and CP4 also inhibited LysRS- CA or WM CA-CTD interaction with an IC50 value of 1 µM. Furthermore, preliminary chemical-shift perturbation NMR analyses suggest that CP 4 binds to a site proximal to helix 4 of the CA-CTD, which is the known site of LysRS interaction.

Keywords: Lysyl tRNA Synthetase, HIV-1, Capsid