The SOCS-box of HIV-1 Vif interacts with ElonginBC by induced-folding to recruit its Cul5-containing ubiquitin ligase complex.
The HIV-1 viral infectivity factor (Vif) protein recruits an E3 ubiquitin ligase complex, comprising the cellular proteins elongin B and C (EloBC), cullin 5 (Cul5) and RING-box 2 (Rbx2), to the anti-viral proteins APOBEC3G (A3G) and APOBEC3F (A3F) and induces their polyubiquitination and proteasomal degradation. In this study, we used purified proteins and direct in vitro binding assays, isothermal titration calorimetry and NMR spectroscopy to describe the molecular mechanism for assembly of the Vif-EloBC ternary complex. We demonstrate that Vif binds to EloBC in two locations, and that both interactions induce structural changes in the SOCS box of Vif as well as EloBC. In particular, in addition to the previously established binding of Vif's BC box to EloC, we report a novel interaction between the conserved Pro-Pro-Leu-Pro motif of Vif and the C-terminal domain of EloB. Using cell-based assays, we further show that this interaction is necessary for the formation of a functional ligase complex, thus establishing a role of this motif. We conclude that HIV-1 Vif engages EloBC via an induced-folding mechanism that does not require additional co-factors, and speculate that these features distinguish Vif from other EloBC specificity factors such as cellular SOCS proteins, and may enhance the prospects of obtaining therapeutic inhibitors of Vif function.
Department of Infectious Diseases, King's College London School of Medicine, London, United Kingdom.
Julien R C
JR
Bergeron
Hendrik
H
Huthoff
Dennis A
DA
Veselkov
Rebecca L
RL
Beavil
Peter J
PJ
Simpson
Stephen J
SJ
Matthews
Michael H
MH
Malim
Mark R
MR
Sanderson
D000595:Amino Acid Sequence; D044842:Cullin Proteins / Q000737:chemistry / Q000378:metabolism*; D000075065:Elongin; D015658:HIV Infections / Q000378:metabolism / Q000473:pathology; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D017510:Protein Folding; D050826:Suppressor of Cytokine Signaling Proteins / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination; D054320:vif Gene Products, Human Immunodeficiency Virus / Q000737:chemistry / Q000378:metabolism*
C496871:CUL5 protein, human; D044842:Cullin Proteins; C000616396:ELOB protein, human; C000616395:ELOC protein, human; D000075065:Elongin; D050826:Suppressor of Cytokine Signaling Proteins; D014157:Transcription Factors; D054320:vif Gene Products, Human Immunodeficiency Virus; D044767:Ubiquitin-Protein Ligases