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Integrative Modeling of Macromolecular Assemblies from Low to Near-Atomic Resolution.

Xu X, Yan C, Wohlhueter R, Ivanov I - Comput Struct Biotechnol J (2015)

Bottom Line: By systematically combining various sources of structural, biochemical and biophysical information, integrative modeling approaches aim to provide a unified structural description of such assemblies, starting from high-resolution structures of the individual components and integrating all available information from low-resolution experimental methods.Second, we describe hybrid molecular dynamics, Rosetta Monte-Carlo and minimum ensemble search (MES) methods that can be used to incorporate SAXS into pseudoatomic structural models.We present concise descriptions of the two methods and their most popular alternatives, along with select illustrative applications to protein/nucleic acid assemblies involved in DNA replication and repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Center for Diagnostics and Therapeutics, Georgia State University, Atlanta, GA 30302, USA.

ABSTRACT
While conventional high-resolution techniques in structural biology are challenged by the size and flexibility of many biological assemblies, recent advances in low-resolution techniques such as cryo-electron microscopy (cryo-EM) and small angle X-ray scattering (SAXS) have opened up new avenues to define the structures of such assemblies. By systematically combining various sources of structural, biochemical and biophysical information, integrative modeling approaches aim to provide a unified structural description of such assemblies, starting from high-resolution structures of the individual components and integrating all available information from low-resolution experimental methods. In this review, we describe integrative modeling approaches, which use complementary data from either cryo-EM or SAXS. Specifically, we focus on the popular molecular dynamics flexible fitting (MDFF) method, which has been widely used for flexible fitting into cryo-EM maps. Second, we describe hybrid molecular dynamics, Rosetta Monte-Carlo and minimum ensemble search (MES) methods that can be used to incorporate SAXS into pseudoatomic structural models. We present concise descriptions of the two methods and their most popular alternatives, along with select illustrative applications to protein/nucleic acid assemblies involved in DNA replication and repair.

No MeSH data available.


Single-particle EM analysis shows how FEN1 interacts flexibly with 9-1-1 and adopts a fixed position on 9Δ-1-1 in the presence of the DNA substrate. (A) Representative reference-free 2D class averages (top and side views) for the 9-1-1/FEN1 binary complex are compared with those corresponding to the 9Δ-1-1/FEN1/DNA ternary complex. Top and side views of the 9-1-1 complex are shown. (B) Side and top views of the 9Δ-1-1/FEN1/DNA 3D reconstruction. (C) MDFF flexible fitting of the 9Δ-1-1/FEN1/DNA complex into the 3D map of the ternary complex.
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f0010: Single-particle EM analysis shows how FEN1 interacts flexibly with 9-1-1 and adopts a fixed position on 9Δ-1-1 in the presence of the DNA substrate. (A) Representative reference-free 2D class averages (top and side views) for the 9-1-1/FEN1 binary complex are compared with those corresponding to the 9Δ-1-1/FEN1/DNA ternary complex. Top and side views of the 9-1-1 complex are shown. (B) Side and top views of the 9Δ-1-1/FEN1/DNA 3D reconstruction. (C) MDFF flexible fitting of the 9Δ-1-1/FEN1/DNA complex into the 3D map of the ternary complex.

Mentions: Using single-particle EM, the structural features of the binary complexes of 9-1-1/FEN1 and the ternary complex of 9-1-1/FEN1/DNA were revealed by the reference-free 2D class averages as shown in Fig. 2A. The computational model of 9-1-1/FEN1/DNA was then filtered at 20 Å to assign relative orientations to the different experimental views of the assembly. A final 3D reconstruction of the 9-1-1/FEN1/DNA complex at a resolution of 18 Å was then obtained using 3D refinement with iterative projection matching [84,85] (Fig. 2B). The atomistic model was then flexibly fitted into the EM map using MDFF. Due to the difficulty in visualizing DNA density with negative staining, the DNA was not included in the MDFF process. In the end, the fitted atomic model of 9-1-1/FEN1/DNA left fewer than 300 atoms outside of the EM map (at a threshold of 3.6), showing an excellent agreement with the map (Fig. 2C). It is worth noting that the EM map supports the observation that FEN1 is tilted toward the Rad1 subunit in the computational model.


Integrative Modeling of Macromolecular Assemblies from Low to Near-Atomic Resolution.

Xu X, Yan C, Wohlhueter R, Ivanov I - Comput Struct Biotechnol J (2015)

Single-particle EM analysis shows how FEN1 interacts flexibly with 9-1-1 and adopts a fixed position on 9Δ-1-1 in the presence of the DNA substrate. (A) Representative reference-free 2D class averages (top and side views) for the 9-1-1/FEN1 binary complex are compared with those corresponding to the 9Δ-1-1/FEN1/DNA ternary complex. Top and side views of the 9-1-1 complex are shown. (B) Side and top views of the 9Δ-1-1/FEN1/DNA 3D reconstruction. (C) MDFF flexible fitting of the 9Δ-1-1/FEN1/DNA complex into the 3D map of the ternary complex.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588362&req=5

f0010: Single-particle EM analysis shows how FEN1 interacts flexibly with 9-1-1 and adopts a fixed position on 9Δ-1-1 in the presence of the DNA substrate. (A) Representative reference-free 2D class averages (top and side views) for the 9-1-1/FEN1 binary complex are compared with those corresponding to the 9Δ-1-1/FEN1/DNA ternary complex. Top and side views of the 9-1-1 complex are shown. (B) Side and top views of the 9Δ-1-1/FEN1/DNA 3D reconstruction. (C) MDFF flexible fitting of the 9Δ-1-1/FEN1/DNA complex into the 3D map of the ternary complex.
Mentions: Using single-particle EM, the structural features of the binary complexes of 9-1-1/FEN1 and the ternary complex of 9-1-1/FEN1/DNA were revealed by the reference-free 2D class averages as shown in Fig. 2A. The computational model of 9-1-1/FEN1/DNA was then filtered at 20 Å to assign relative orientations to the different experimental views of the assembly. A final 3D reconstruction of the 9-1-1/FEN1/DNA complex at a resolution of 18 Å was then obtained using 3D refinement with iterative projection matching [84,85] (Fig. 2B). The atomistic model was then flexibly fitted into the EM map using MDFF. Due to the difficulty in visualizing DNA density with negative staining, the DNA was not included in the MDFF process. In the end, the fitted atomic model of 9-1-1/FEN1/DNA left fewer than 300 atoms outside of the EM map (at a threshold of 3.6), showing an excellent agreement with the map (Fig. 2C). It is worth noting that the EM map supports the observation that FEN1 is tilted toward the Rad1 subunit in the computational model.

Bottom Line: By systematically combining various sources of structural, biochemical and biophysical information, integrative modeling approaches aim to provide a unified structural description of such assemblies, starting from high-resolution structures of the individual components and integrating all available information from low-resolution experimental methods.Second, we describe hybrid molecular dynamics, Rosetta Monte-Carlo and minimum ensemble search (MES) methods that can be used to incorporate SAXS into pseudoatomic structural models.We present concise descriptions of the two methods and their most popular alternatives, along with select illustrative applications to protein/nucleic acid assemblies involved in DNA replication and repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Center for Diagnostics and Therapeutics, Georgia State University, Atlanta, GA 30302, USA.

ABSTRACT
While conventional high-resolution techniques in structural biology are challenged by the size and flexibility of many biological assemblies, recent advances in low-resolution techniques such as cryo-electron microscopy (cryo-EM) and small angle X-ray scattering (SAXS) have opened up new avenues to define the structures of such assemblies. By systematically combining various sources of structural, biochemical and biophysical information, integrative modeling approaches aim to provide a unified structural description of such assemblies, starting from high-resolution structures of the individual components and integrating all available information from low-resolution experimental methods. In this review, we describe integrative modeling approaches, which use complementary data from either cryo-EM or SAXS. Specifically, we focus on the popular molecular dynamics flexible fitting (MDFF) method, which has been widely used for flexible fitting into cryo-EM maps. Second, we describe hybrid molecular dynamics, Rosetta Monte-Carlo and minimum ensemble search (MES) methods that can be used to incorporate SAXS into pseudoatomic structural models. We present concise descriptions of the two methods and their most popular alternatives, along with select illustrative applications to protein/nucleic acid assemblies involved in DNA replication and repair.

No MeSH data available.