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Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration

Integration of retroviral DNA into the host genome involves the formation of integrase (IN)–DNA complexes termed intasomes. Further characterization of these complexes is needed to understand their assembly process. Here, we report the single-particle cryo-EM structure of the Rous sarcoma virus (RSV...

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Autores principales: Bera, Sibes, Shi, Ke, Aihara, Hideki, Grandgenett, Duane P., Pandey, Krishan K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Biochemistry and Molecular Biology 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10209032/
https://www.ncbi.nlm.nih.gov/pubmed/37084813
http://dx.doi.org/10.1016/j.jbc.2023.104730
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author Bera, Sibes
Shi, Ke
Aihara, Hideki
Grandgenett, Duane P.
Pandey, Krishan K.
author_facet Bera, Sibes
Shi, Ke
Aihara, Hideki
Grandgenett, Duane P.
Pandey, Krishan K.
author_sort Bera, Sibes
collection PubMed
description Integration of retroviral DNA into the host genome involves the formation of integrase (IN)–DNA complexes termed intasomes. Further characterization of these complexes is needed to understand their assembly process. Here, we report the single-particle cryo-EM structure of the Rous sarcoma virus (RSV) strand transfer complex (STC) intasome produced with IN and a preassembled viral/target DNA substrate at 3.36 Å resolution. The conserved intasome core region consisting of IN subunits contributing active sites interacting with viral/target DNA has a resolution of 3 Å. Our structure demonstrated the flexibility of the distal IN subunits relative to the IN subunits in the conserved intasome core, similar to results previously shown with the RSV octameric cleaved synaptic complex intasome produced with IN and viral DNA only. An extensive analysis of higher resolution STC structure helped in the identification of nucleoprotein interactions important for intasome assembly. Using structure-function studies, we determined the mechanisms of several IN–DNA interactions critical for assembly of both RSV intasomes. We determined the role of IN residues R244, Y246, and S124 in cleaved synaptic complex and STC intasome assemblies and their catalytic activities, demonstrating differential effects. Taken together, these studies advance our understanding of different RSV intasome structures and molecular determinants involved in their assembly.
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spelling pubmed-102090322023-05-26 Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration Bera, Sibes Shi, Ke Aihara, Hideki Grandgenett, Duane P. Pandey, Krishan K. J Biol Chem Research Article Integration of retroviral DNA into the host genome involves the formation of integrase (IN)–DNA complexes termed intasomes. Further characterization of these complexes is needed to understand their assembly process. Here, we report the single-particle cryo-EM structure of the Rous sarcoma virus (RSV) strand transfer complex (STC) intasome produced with IN and a preassembled viral/target DNA substrate at 3.36 Å resolution. The conserved intasome core region consisting of IN subunits contributing active sites interacting with viral/target DNA has a resolution of 3 Å. Our structure demonstrated the flexibility of the distal IN subunits relative to the IN subunits in the conserved intasome core, similar to results previously shown with the RSV octameric cleaved synaptic complex intasome produced with IN and viral DNA only. An extensive analysis of higher resolution STC structure helped in the identification of nucleoprotein interactions important for intasome assembly. Using structure-function studies, we determined the mechanisms of several IN–DNA interactions critical for assembly of both RSV intasomes. We determined the role of IN residues R244, Y246, and S124 in cleaved synaptic complex and STC intasome assemblies and their catalytic activities, demonstrating differential effects. Taken together, these studies advance our understanding of different RSV intasome structures and molecular determinants involved in their assembly. American Society for Biochemistry and Molecular Biology 2023-04-20 /pmc/articles/PMC10209032/ /pubmed/37084813 http://dx.doi.org/10.1016/j.jbc.2023.104730 Text en © 2023 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Research Article
Bera, Sibes
Shi, Ke
Aihara, Hideki
Grandgenett, Duane P.
Pandey, Krishan K.
Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title_full Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title_fullStr Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title_full_unstemmed Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title_short Molecular determinants for Rous sarcoma virus intasome assemblies involved in retroviral integration
title_sort molecular determinants for rous sarcoma virus intasome assemblies involved in retroviral integration
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10209032/
https://www.ncbi.nlm.nih.gov/pubmed/37084813
http://dx.doi.org/10.1016/j.jbc.2023.104730
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