Dataset Information

Tubular crystals and helical arrays: structural determination of HIV-1 capsid assemblies using iterative helical real-space reconstruction.

ABSTRACT: Helical structures are important in many different life forms and are well-suited for structural studies by cryo-EM. A unique feature of helical objects is that a single projection image contains all the views needed to perform a three-dimensional (3D) crystallographic reconstruction. Here, we use HIV-1 capsid assemblies to illustrate the detailed approaches to obtain 3D density maps from helical objects. Mature HIV-1 particles contain a conical- or tubular-shaped capsid that encloses the viral RNA genome and performs essential functions in the virus life cycle. The capsid is composed of capsid protein (CA) oligomers which are helically arranged on the surface. The N-terminal domain (NTD) of CA is connected to its C-terminal domain (CTD) through a flexible hinge. Structural analysis of two- and three-dimensional crystals provided molecular models of the capsid protein (CA) and its oligomer forms. We determined the 3D density map of helically assembled HIV-1 CA hexamers at 16 Å resolution using an iterative helical real-space reconstruction method. Docking of atomic models of CA-NTD and CA-CTD dimer into the electron density map indicated that the CTD dimer interface is retained in the assembled CA. Furthermore, molecular docking revealed an additional, novel CTD trimer interface.

SUBMITTER: Zhang P

PROVIDER: S-EPMC3748364 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Tubular crystals and helical arrays: structural determination of HIV-1 capsid assemblies using iterative helical real-space reconstruction.

Zhang Peijun P Meng Xin X Zhao Gongpu G

Methods in molecular biology (Clifton, N.J.) 20130101

Helical structures are important in many different life forms and are well-suited for structural studies by cryo-EM. A unique feature of helical objects is that a single projection image contains all the views needed to perform a three-dimensional (3D) crystallographic reconstruction. Here, we use HIV-1 capsid assemblies to illustrate the detailed approaches to obtain 3D density maps from helical objects. Mature HIV-1 particles contain a conical- or tubular-shaped capsid that encloses the viral ...[more]

PMID: 23132072

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Project description:The HIV-1 capsid protein (CA) forms the capsid shell that encloses RNA within a mature HIV-1 virion. Previous studies by electron microscopy have shown that the capsid shell is primarily a triangular lattice of CA hexamers, with variable curvature that destroys the ideal symmetry of a planar lattice. The mature CA lattice depends on CA dimerization, which occurs through interactions between helix 9 segments of the C-terminal domain (CTD) of CA. Several high-resolution structures of the CTD-CTD dimerization interface have been reported, based on X-ray crystallography and multidimensional solution nuclear magnetic resonance (NMR), with significant differences in amino acid side chain conformations and helix 9-helix 9 orientations. In a structural model for tubular CA assemblies based on cryogenic electron microscopy (cryoEM) [Zhao et al. Nature, 2013, 497, 643-646], the dimerization interface is substantially disordered. The dimerization interface structure in noncrystalline CA assemblies and the extent to which this interface is structurally ordered within a curved lattice have therefore been unclear. Here we describe solid state NMR measurements on the dimerization interface in tubular CA assemblies, which contain the curved triangular lattice of a mature virion, including quantitative measurements of intermolecular and intramolecular distances using dipolar recoupling techniques, solid state NMR chemical shifts, and long-range side chain-side chain contacts. When combined with restraints on the distance and orientation between helix 9 segments from the cryoEM study, the solid state NMR data lead to a unique high-resolution structure for the dimerization interface in the noncrystalline lattice of CA tubes. These results demonstrate that CA lattice curvature is not dependent on disorder or variability in the dimerization interface. This work also demonstrates the feasibility of local structure determination within large noncrystalline assemblies formed by high-molecular-weight proteins, using modern solid state NMR methods.

Dataset Information

Tubular crystals and helical arrays: structural determination of HIV-1 capsid assemblies using iterative helical real-space reconstruction.

Publications

Tubular crystals and helical arrays: structural determination of HIV-1 capsid assemblies using iterative helical real-space reconstruction.

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