Co-crystal structures of HIV TAR RNA bound to lab-evolved proteins show key roles for arginine relevant to the design of cyclic peptide TAR inhibitors.
J Biol Chem
; 295(49): 16470-16486, 2020 12 04.
Article
em En
| MEDLINE
| ID: mdl-33051202
ABSTRACT
RNA-protein interfaces control key replication events during the HIV-1 life cycle. The viral trans-activator of transcription (Tat) protein uses an archetypal arginine-rich motif (ARM) to recruit the host positive transcription elongation factor b (pTEFb) complex onto the viral trans-activation response (TAR) RNA, leading to activation of HIV transcription. Efforts to block this interaction have stimulated production of biologics designed to disrupt this essential RNA-protein interface. Here, we present four co-crystal structures of lab-evolved TAR-binding proteins (TBPs) in complex with HIV-1 TAR. Our results reveal that high-affinity binding requires a distinct sequence and spacing of arginines within a specific ß2-ß3 hairpin loop that arose during selection. Although loops with as many as five arginines were analyzed, only three arginines could bind simultaneously with major-groove guanines. Amino acids that promote backbone interactions within the ß2-ß3 loop were also observed to be important for high-affinity interactions. Based on structural and affinity analyses, we designed two cyclic peptide mimics of the TAR-binding ß2-ß3 loop sequences present in two high-affinity TBPs (KD values of 4.2 ± 0.3 and 3.0 ± 0.3 nm). Our efforts yielded low-molecular weight compounds that bind TAR with low micromolar affinity (KD values ranging from 3.6 to 22 µm). Significantly, one cyclic compound within this series blocked binding of the Tat-ARM peptide to TAR in solution assays, whereas its linear counterpart did not. Overall, this work provides insight into protein-mediated TAR recognition and lays the ground for the development of cyclic peptide inhibitors of a vital HIV-1 RNA-protein interaction.
Palavras-chave
HIV TAR; HIV Tat; RNA structure; RNA-binding protein; RNA-protein interaction; RNA-protein interactions; X-ray crystallography; arginine-rich domain; cyclic peptide; cyclic peptide inhibitor; drug discovery; human immunodeficiency virus (HIV); isothermal titration calorimetry; isothermal titration calorimetry (ITC); peptide chemical synthesis; surface plasmon resonance; surface plasmon resonance (SPR)
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Peptídeos Cíclicos
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Arginina
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RNA Viral
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Repetição Terminal Longa de HIV
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HIV-1
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Proteína de Ligação a TATA-Box
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article