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Lys63-linked ubiquitin chain adopts multiple conformational states for specific target recognition.

Liu Z, Gong Z, Jiang WX, Yang J, Zhu WK, Guo DC, Zhang WP, Liu ML, Tang C - Elife (2015)

Bottom Line: Free or bound to ligands, polyubiquitins are found in different arrangements of ubiquitin subunits.A point mutation that shifts the equilibrium between the different states modulates the binding affinities towards K63-Ub2 ligands.This conformational selection mechanism at the quaternary level may be used by polyubiquitins of different lengths and linkages for target recognition.

View Article: PubMed Central - PubMed

Affiliation: CAS Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic Molecular Physics, Wuhan Institute of Physics and Mathematics of the Chinese Academy of Sciences, Wuhan, China.

ABSTRACT
A polyubiquitin comprises multiple covalently linked ubiquitins and recognizes myriad targets. Free or bound to ligands, polyubiquitins are found in different arrangements of ubiquitin subunits. To understand the structural basis for polyubiquitin quaternary plasticity and to explore the target recognition mechanism, we characterize the conformational space of Lys63-linked diubiquitin (K63-Ub2). Refining against inter-subunit paramagnetic NMR data, we show that free K63-Ub2 exists as a dynamic ensemble comprising multiple closed and open quaternary states. The quaternary dynamics enables K63-Ub2 to be specifically recognized in a variety of signaling pathways. When binding to a target protein, one of the preexisting quaternary states is selected and stabilized. A point mutation that shifts the equilibrium between the different states modulates the binding affinities towards K63-Ub2 ligands. This conformational selection mechanism at the quaternary level may be used by polyubiquitins of different lengths and linkages for target recognition.

No MeSH data available.


Definition of the spherical coordinate system.The origin is set at the center-of-mass of the distal unit, with north pole indicated. With the distal unit fixed, the polar angle of the proximal unit defines the relative orientation of the vector connecting the centers-of-mass of the distal and proximal units (dashed lines).DOI:http://dx.doi.org/10.7554/eLife.05767.018
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fig4s1: Definition of the spherical coordinate system.The origin is set at the center-of-mass of the distal unit, with north pole indicated. With the distal unit fixed, the polar angle of the proximal unit defines the relative orientation of the vector connecting the centers-of-mass of the distal and proximal units (dashed lines).DOI:http://dx.doi.org/10.7554/eLife.05767.018

Mentions: To better visualize the ensemble structure of K63-Ub2 in the closed state, we projected the position of the proximal unit relative to the distal unit using spherical coordinates (Figure 4—figure supplement 1). Upon reducing the dimensionality, we found that the closed-state structures exist in two clusters, namely C1 and C2 (Figure 4A). For each four-conformer structure, one of the conformers falls into C2, while the other three are in C1. The proximal unit of ligand-free K63-Ub2 utilizes distinct interfaces to interact with the distal unit in C1 and C2 states (Figure 4B,C), affording buried solvent-accessible surface areas of 283.9 ± 139.7 Å2 and 200.5 ± 59.6 Å2, respectively. Significantly, the crystal structures of K63-Ub2 in complex with the ZnF4 domain of A20 (Bosanac et al., 2010) and with the NZF domain of TAB2 or TAB3 (Kulathu et al., 2009; Sato et al., 2009b) are found within or near the C1 and C2 clusters, respectively (Figure 4A). The root-mean-square difference (RMSD) between the conformers in C1 and the ZnF4-bound structure of K63-Ub2 is as small as 3.93 Å (Figure 4B), while the RMSD between C2 conformers and the NZF-bound structure is as small as 1.68 Å (Figure 4C). We predicted the inter-subunit PREs for two known complex structures in closed states (Figure 4—figure supplement 2A,B). Linearly combining the two sets of PREs at a 3:1 ratio and 70% total population, the resulting PREs agree well with the experimental values, although some details differ (Figure 4—figure supplement 2C). On the other hand, the paired-distance distribution profiles computed for A20 ZnF4 and TAB2/TAB3 NZF complexed K63-Ub2 (with bound ligand removed) display narrower distributions compared to those computed for the open-state structures or to the experimental data (Figure 2A).10.7554/eLife.05767.017Figure 4.Ensemble structure of K63-Ub2 in closed state.


Lys63-linked ubiquitin chain adopts multiple conformational states for specific target recognition.

Liu Z, Gong Z, Jiang WX, Yang J, Zhu WK, Guo DC, Zhang WP, Liu ML, Tang C - Elife (2015)

Definition of the spherical coordinate system.The origin is set at the center-of-mass of the distal unit, with north pole indicated. With the distal unit fixed, the polar angle of the proximal unit defines the relative orientation of the vector connecting the centers-of-mass of the distal and proximal units (dashed lines).DOI:http://dx.doi.org/10.7554/eLife.05767.018
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4507786&req=5

fig4s1: Definition of the spherical coordinate system.The origin is set at the center-of-mass of the distal unit, with north pole indicated. With the distal unit fixed, the polar angle of the proximal unit defines the relative orientation of the vector connecting the centers-of-mass of the distal and proximal units (dashed lines).DOI:http://dx.doi.org/10.7554/eLife.05767.018
Mentions: To better visualize the ensemble structure of K63-Ub2 in the closed state, we projected the position of the proximal unit relative to the distal unit using spherical coordinates (Figure 4—figure supplement 1). Upon reducing the dimensionality, we found that the closed-state structures exist in two clusters, namely C1 and C2 (Figure 4A). For each four-conformer structure, one of the conformers falls into C2, while the other three are in C1. The proximal unit of ligand-free K63-Ub2 utilizes distinct interfaces to interact with the distal unit in C1 and C2 states (Figure 4B,C), affording buried solvent-accessible surface areas of 283.9 ± 139.7 Å2 and 200.5 ± 59.6 Å2, respectively. Significantly, the crystal structures of K63-Ub2 in complex with the ZnF4 domain of A20 (Bosanac et al., 2010) and with the NZF domain of TAB2 or TAB3 (Kulathu et al., 2009; Sato et al., 2009b) are found within or near the C1 and C2 clusters, respectively (Figure 4A). The root-mean-square difference (RMSD) between the conformers in C1 and the ZnF4-bound structure of K63-Ub2 is as small as 3.93 Å (Figure 4B), while the RMSD between C2 conformers and the NZF-bound structure is as small as 1.68 Å (Figure 4C). We predicted the inter-subunit PREs for two known complex structures in closed states (Figure 4—figure supplement 2A,B). Linearly combining the two sets of PREs at a 3:1 ratio and 70% total population, the resulting PREs agree well with the experimental values, although some details differ (Figure 4—figure supplement 2C). On the other hand, the paired-distance distribution profiles computed for A20 ZnF4 and TAB2/TAB3 NZF complexed K63-Ub2 (with bound ligand removed) display narrower distributions compared to those computed for the open-state structures or to the experimental data (Figure 2A).10.7554/eLife.05767.017Figure 4.Ensemble structure of K63-Ub2 in closed state.

Bottom Line: Free or bound to ligands, polyubiquitins are found in different arrangements of ubiquitin subunits.A point mutation that shifts the equilibrium between the different states modulates the binding affinities towards K63-Ub2 ligands.This conformational selection mechanism at the quaternary level may be used by polyubiquitins of different lengths and linkages for target recognition.

View Article: PubMed Central - PubMed

Affiliation: CAS Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic Molecular Physics, Wuhan Institute of Physics and Mathematics of the Chinese Academy of Sciences, Wuhan, China.

ABSTRACT
A polyubiquitin comprises multiple covalently linked ubiquitins and recognizes myriad targets. Free or bound to ligands, polyubiquitins are found in different arrangements of ubiquitin subunits. To understand the structural basis for polyubiquitin quaternary plasticity and to explore the target recognition mechanism, we characterize the conformational space of Lys63-linked diubiquitin (K63-Ub2). Refining against inter-subunit paramagnetic NMR data, we show that free K63-Ub2 exists as a dynamic ensemble comprising multiple closed and open quaternary states. The quaternary dynamics enables K63-Ub2 to be specifically recognized in a variety of signaling pathways. When binding to a target protein, one of the preexisting quaternary states is selected and stabilized. A point mutation that shifts the equilibrium between the different states modulates the binding affinities towards K63-Ub2 ligands. This conformational selection mechanism at the quaternary level may be used by polyubiquitins of different lengths and linkages for target recognition.

No MeSH data available.