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Self-assembly of Rous Sarcoma Virus capsid protein, probed by Solid-state NMR and TEM

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Date Issued:
2016
Abstract/Description:
The retroviral capsid protein (CA) is derived from the cleavage of Gag polyprotein during the maturation process, and self-assembles into a polymorphic fullerene-like shell encasing the viral genome materials. The orthoretroviral CAs, for instance, Human Immunodeficiency Virus (HIV) and Rous Sarcoma Virus (RSV) CA, has little similarity in their sequence composition but a common 3D structure. They form distinct capsid assembly in vivo and a range of similar assemblies in vitro. Due to the substantial polymorphism, such assemblies are not amenable for conventional structural biology techniques such as X-ray diffraction crystallography and cryo-electron microscopy (cryo-EM). Solid-state NMR spectroscopy is the optimal platform to study these CA assemblies to attain site-specific structural and dynamic information. However, it is challenging to make signal assignments for such non-crystalline and large biomolecules as retroviral CA assemblies. In this study, we were to elucidate the assembly mechanism of retroviral capsids by applying the state-of-art solid-state NMR techniques on the RSV CA assembly system and establishing an atomistic resolution structural model. The RSV CA is the second most studied protein among the retroviral family after HIV that causes AIDS (acquired immune deficiency syndrome), but there is no atomistic model for RSV CA assemblies available.In this study, we showed that highly uniform tubular RSV CA assembly can be prepared. Screened by TEM, our tubular assembly showed sharp 6-fold symmetry under diffraction, illustrating the quasi-crystalline character. Subsequently we acquired a series of solid-state NMRivspectra for tubular RSV CA assembly, and completed chemical shift signal assignments with samples of various isotope labeling. Then, combining cryo-EM electron density map of tubular assembly of RSV CA with the secondary structures derived from solid-state NMR, we established an atomistic resolution structure model. In this model, we identified the residue-specific assembly interfaces. Interestingly, our model revealed the structural re-arrangements upon the assembly and suggested that the tubular assembly of RSV CA may take a different assembly pathway from that of HIV capsid.
Title: Self-assembly of Rous Sarcoma Virus capsid protein, probed by Solid-state NMR and TEM.
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Name(s): Jeon, Jaekyun, Author
Chen, Bo, Committee Chair
Tatulian, Suren, Committee CoChair
Schulte, Alfons, Committee Member
Cole, Alexander, Committee Member
University of Central Florida, Degree Grantor
Type of Resource: text
Date Issued: 2016
Publisher: University of Central Florida
Language(s): English
Abstract/Description: The retroviral capsid protein (CA) is derived from the cleavage of Gag polyprotein during the maturation process, and self-assembles into a polymorphic fullerene-like shell encasing the viral genome materials. The orthoretroviral CAs, for instance, Human Immunodeficiency Virus (HIV) and Rous Sarcoma Virus (RSV) CA, has little similarity in their sequence composition but a common 3D structure. They form distinct capsid assembly in vivo and a range of similar assemblies in vitro. Due to the substantial polymorphism, such assemblies are not amenable for conventional structural biology techniques such as X-ray diffraction crystallography and cryo-electron microscopy (cryo-EM). Solid-state NMR spectroscopy is the optimal platform to study these CA assemblies to attain site-specific structural and dynamic information. However, it is challenging to make signal assignments for such non-crystalline and large biomolecules as retroviral CA assemblies. In this study, we were to elucidate the assembly mechanism of retroviral capsids by applying the state-of-art solid-state NMR techniques on the RSV CA assembly system and establishing an atomistic resolution structural model. The RSV CA is the second most studied protein among the retroviral family after HIV that causes AIDS (acquired immune deficiency syndrome), but there is no atomistic model for RSV CA assemblies available.In this study, we showed that highly uniform tubular RSV CA assembly can be prepared. Screened by TEM, our tubular assembly showed sharp 6-fold symmetry under diffraction, illustrating the quasi-crystalline character. Subsequently we acquired a series of solid-state NMRivspectra for tubular RSV CA assembly, and completed chemical shift signal assignments with samples of various isotope labeling. Then, combining cryo-EM electron density map of tubular assembly of RSV CA with the secondary structures derived from solid-state NMR, we established an atomistic resolution structure model. In this model, we identified the residue-specific assembly interfaces. Interestingly, our model revealed the structural re-arrangements upon the assembly and suggested that the tubular assembly of RSV CA may take a different assembly pathway from that of HIV capsid.
Identifier: CFE0006332 (IID), ucf:51572 (fedora)
Note(s): 2016-08-01
Ph.D.
Sciences, Physics
Doctoral
This record was generated from author submitted information.
Subject(s): retroviral capsid -- RSV capsid -- solid-state NMR -- TEM
Persistent Link to This Record: http://purl.flvc.org/ucf/fd/CFE0006332
Restrictions on Access: campus 2017-08-15
Host Institution: UCF

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