Biological properties of reverse ankyrin engineered for dimer construction to enhance HIV-1 capsid interaction.
Asian Pac J Allergy Immunol
; 2022 May 22.
Article
em En
| MEDLINE
| ID: mdl-35598193
ABSTRACT
BACKGROUND:
Assembly and budding in the late-stage of human immunodeficiency virus type 1 (HIV-1) production rely on Gag protein polymerization at the inner leaflet of the plasma membrane. We previously generated a monomeric ankyrin repeat protein (Ank1D4) that specifically interacts with capsid protein (CAp24) of HIV-1, however this protein had modest binding affinity.OBJECTIVE:
This study aimed to improve the avidity of Ank1D4 by generating two Ank1D4 dimers (Ank1D4NC-NC) and its inverted form (Ank1D4NC-CN), with each domain connected by a flexible (G4S)4 linker peptide.METHODS:
Binding properties of monomeric and dimeric Ank1D4 was performed by capture enzyme-linked immunosorbent assay (ELISA). Sandwich ELISA was used to examine bifunctional module of dimeric Ank1D4. Ank1D4NC-NC and Ank1D4NC-CN were evaluated using bio-layer interferometry (BLI), compared to monomeric Ank1D4.RESULTS:
Similar binding surfaces were observed in both dimers which was comparable with monomeric Ank1D4. The interaction of Ank1D4NC-CN with CAp24 was significantly greater than that of Ank1D4NC-NC and Ank1D4 by capture ELISA. Ank1D4NC-CN also exhibited bifunctionality using a sandwich ELISA. The KD of Ank1D4NC-CN, Ank1D4NC-NC and monomeric Ank1D4 was 3.5 nM, 53.7 nM, and 126.2 nM, respectively using bio-layer interferometry analysis.CONCLUSIONS:
This study provides a strategy for increasing Ank1D4 avidity through the construction of novel inverted dimers with a flexible linker. Ank1D4NC-CN may provide an alternative treatment strategy for inhibiting HIV-1 replication.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Asian Pac J Allergy Immunol
Ano de publicação:
2022
Tipo de documento:
Article
País de afiliação:
Tailândia