Journal Article

Characterization of the Interaction of Full-Length HIV-1 Vif Protein with its Key Regulator CBFβ and CRL5 E3 Ubiquitin Ligase Components

Zhou, Xiaohong, Liu, Yayan, Yu, Xiao-Fang, Evans, Sean L., Han, Xue

2012

Overview

Human immunodeficiency virus-1 (HIV-1) viral infectivity factor (Vif) is essential for viral replication because of its ability to eliminate the host's antiviral response to HIV-1 that is mediated by the APOBEC3 family of cellular cytidine deaminases. Vif targets these proteins, including APOBEC3G, for polyubiquitination and subsequent proteasome-mediated degradation via the formation of a Cullin5-ElonginB/C-based E3 ubiquitin ligase. Determining how the cellular components of this E3 ligase complex interact with Vif is critical to the intelligent design of new antiviral drugs. However, structural studies of Vif, both alone and in complex with cellular partners, have been hampered by an inability to express soluble full-length Vif protein. Here we demonstrate that a newly identified host regulator of Vif, core-binding factor-beta (CBFβ), interacts directly with Vif, including various isoforms and a truncated form of this regulator. In addition, carboxyl-terminal truncations of Vif lacking the BC-box and cullin box motifs were sufficient for CBFβ interaction. Furthermore, association of Vif with CBFβ, alone or in combination with Elongin B/C (EloB/C), greatly increased the solubility of full-length Vif. Finally, a stable complex containing Vif-CBFβ-EloB/C was purified in large quantity and shown to bind purified Cullin5 (Cul5). This efficient strategy for purifying Vif-Cul5-CBFβ-EloB/C complexes will facilitate future structural and biochemical studies of Vif function and may provide the basis for useful screening approaches for identifying novel anti-HIV drug candidates.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Acquired immune deficiency syndrome / Aggregates / AIDS / Amino Acid Sequence / Animals / Antiviral agents / Biology / Cancer / Cellular structure / Cloning / Core Binding Factor beta Subunit - genetics / Core Binding Factor beta Subunit - metabolism / Cullin / Cullin Proteins - metabolism / Deoxyribonucleic acid / DNA / Drug development / Drugs / Elongin / Gene Expression / HIV / Human immunodeficiency virus / Humans / Immunology / Infectivity / Isoforms / Leukemia / Molecular Sequence Data / Multiprotein Complexes - isolation & purification / Multiprotein Complexes - metabolism / Mutation / Proteasomes / Protein Binding / Protein Interaction Domains and Motifs / Protein Stability / Proteins / Public health / Recombinant Fusion Proteins - chemistry / Recombinant Fusion Proteins - genetics / Recombinant Fusion Proteins - metabolism / Scientific imaging / Solubility / Studies / Transcription Factors - genetics / Transcription Factors - metabolism / Ubiquitin / Ubiquitin-protein ligase / vif Gene Products, Human Immunodeficiency Virus - chemistry / vif Gene Products, Human Immunodeficiency Virus - genetics / vif Gene Products, Human Immunodeficiency Virus - metabolism / Vif protein / Virology / Viruses