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Examinations of tRNA Range of Motion Using Simulations of Cryo-EM Microscopy and X-Ray Data.

Caulfield TR, Devkota B, Rollins GC - J Biophys (2011)

Bottom Line: The unbiased molecular dynamics data describes the global conformational changes of tRNA on a sub-microsecond time scale for comparison with steered data.Additionally, the unbiased molecular dynamics data was used to identify putative contacts between tRNA and the ribosome during the accommodation step of translation.We found that the primary contact regions were H71 and H92 of the 50S subunit and ribosomal proteins L14 and L16.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry & Biochemistry, Georgia Institute of Technology, 901 Atlantic Avenue, Atlanta, GA 30332-0230, USA.

ABSTRACT
We examined tRNA flexibility using a combination of steered and unbiased molecular dynamics simulations. Using Maxwell's demon algorithm, molecular dynamics was used to steer X-ray structure data toward that from an alternative state obtained from cryogenic-electron microscopy density maps. Thus, we were able to fit X-ray structures of tRNA onto cryogenic-electron microscopy density maps for hybrid states of tRNA. Additionally, we employed both Maxwell's demon molecular dynamics simulations and unbiased simulation methods to identify possible ribosome-tRNA contact areas where the ribosome may discriminate tRNAs during translation. Herein, we collected >500 ns of simulation data to assess the global range of motion for tRNAs. Biased simulations can be used to steer between known conformational stop points, while unbiased simulations allow for a general testing of conformational space previously unexplored. The unbiased molecular dynamics data describes the global conformational changes of tRNA on a sub-microsecond time scale for comparison with steered data. Additionally, the unbiased molecular dynamics data was used to identify putative contacts between tRNA and the ribosome during the accommodation step of translation. We found that the primary contact regions were H71 and H92 of the 50S subunit and ribosomal proteins L14 and L16.

No MeSH data available.


Cryogenic-electron microscopy fitting using Maxwell's demon Molecular Dynamics for tRNAs A/T, A, P, and P/E is shown.  All tRNA atomic models are shown in ribbons/CPK, while densities are as solid.  (a) A/T-site structure for tRNA fit to density, cross-correlation coefficient (CCC) of 95% between the structures theoretical density and the cryo-EM experimental density (Divergent stereo).  (b) A-site structure for tRNA fit to density with a CCC of 83% (Divergent stereo). (c) P-site structure for tRNA with a CCC of 86% (Divergent stereo).  (d) P/E-site structure with a CCC >90% (Divergent stereo).
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fig4: Cryogenic-electron microscopy fitting using Maxwell's demon Molecular Dynamics for tRNAs A/T, A, P, and P/E is shown. All tRNA atomic models are shown in ribbons/CPK, while densities are as solid. (a) A/T-site structure for tRNA fit to density, cross-correlation coefficient (CCC) of 95% between the structures theoretical density and the cryo-EM experimental density (Divergent stereo). (b) A-site structure for tRNA fit to density with a CCC of 83% (Divergent stereo). (c) P-site structure for tRNA with a CCC of 86% (Divergent stereo). (d) P/E-site structure with a CCC >90% (Divergent stereo).

Mentions: Using MdMD, we obtained pathway transitions from the crystal structure state (1EHZ) to the cryo-EM states A, P, A/T, and P/E using less than 10 nanoseconds of molecular dynamic simulation (Supplementary Movies S4, S6). From these simulations, the ensembles of structures are consistent with the density distributions from the cryo-EM density maps. Figure 4 shows the quality of fit for the four structures. Supplementary Table S2 identifies the cryo-EM density maps of each structure and gives the RMSD of each structure relative to the crystal structure of yeast tRNAPhe (PDB code: 1EHZ) [33]. All-atom root mean-squared deviation (RMSD) analyses between the crystal structure and regions from our molecular dynamics model are <2 Å at these superposed regions, indicating that the fine structure was maintained. Using MdMD, the A-site conformation can be forced rapidly to the kinked conformation required for the A/T state. The kinked structure generated using MdMD is within 1.0 Å RMSD of the original manually modeled structure of the A/T tRNA (Supplementary Movie S6) [13].


Examinations of tRNA Range of Motion Using Simulations of Cryo-EM Microscopy and X-Ray Data.

Caulfield TR, Devkota B, Rollins GC - J Biophys (2011)

Cryogenic-electron microscopy fitting using Maxwell's demon Molecular Dynamics for tRNAs A/T, A, P, and P/E is shown.  All tRNA atomic models are shown in ribbons/CPK, while densities are as solid.  (a) A/T-site structure for tRNA fit to density, cross-correlation coefficient (CCC) of 95% between the structures theoretical density and the cryo-EM experimental density (Divergent stereo).  (b) A-site structure for tRNA fit to density with a CCC of 83% (Divergent stereo). (c) P-site structure for tRNA with a CCC of 86% (Divergent stereo).  (d) P/E-site structure with a CCC >90% (Divergent stereo).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig4: Cryogenic-electron microscopy fitting using Maxwell's demon Molecular Dynamics for tRNAs A/T, A, P, and P/E is shown. All tRNA atomic models are shown in ribbons/CPK, while densities are as solid. (a) A/T-site structure for tRNA fit to density, cross-correlation coefficient (CCC) of 95% between the structures theoretical density and the cryo-EM experimental density (Divergent stereo). (b) A-site structure for tRNA fit to density with a CCC of 83% (Divergent stereo). (c) P-site structure for tRNA with a CCC of 86% (Divergent stereo). (d) P/E-site structure with a CCC >90% (Divergent stereo).
Mentions: Using MdMD, we obtained pathway transitions from the crystal structure state (1EHZ) to the cryo-EM states A, P, A/T, and P/E using less than 10 nanoseconds of molecular dynamic simulation (Supplementary Movies S4, S6). From these simulations, the ensembles of structures are consistent with the density distributions from the cryo-EM density maps. Figure 4 shows the quality of fit for the four structures. Supplementary Table S2 identifies the cryo-EM density maps of each structure and gives the RMSD of each structure relative to the crystal structure of yeast tRNAPhe (PDB code: 1EHZ) [33]. All-atom root mean-squared deviation (RMSD) analyses between the crystal structure and regions from our molecular dynamics model are <2 Å at these superposed regions, indicating that the fine structure was maintained. Using MdMD, the A-site conformation can be forced rapidly to the kinked conformation required for the A/T state. The kinked structure generated using MdMD is within 1.0 Å RMSD of the original manually modeled structure of the A/T tRNA (Supplementary Movie S6) [13].

Bottom Line: The unbiased molecular dynamics data describes the global conformational changes of tRNA on a sub-microsecond time scale for comparison with steered data.Additionally, the unbiased molecular dynamics data was used to identify putative contacts between tRNA and the ribosome during the accommodation step of translation.We found that the primary contact regions were H71 and H92 of the 50S subunit and ribosomal proteins L14 and L16.

View Article: PubMed Central - PubMed

Affiliation: School of Chemistry & Biochemistry, Georgia Institute of Technology, 901 Atlantic Avenue, Atlanta, GA 30332-0230, USA.

ABSTRACT
We examined tRNA flexibility using a combination of steered and unbiased molecular dynamics simulations. Using Maxwell's demon algorithm, molecular dynamics was used to steer X-ray structure data toward that from an alternative state obtained from cryogenic-electron microscopy density maps. Thus, we were able to fit X-ray structures of tRNA onto cryogenic-electron microscopy density maps for hybrid states of tRNA. Additionally, we employed both Maxwell's demon molecular dynamics simulations and unbiased simulation methods to identify possible ribosome-tRNA contact areas where the ribosome may discriminate tRNAs during translation. Herein, we collected >500 ns of simulation data to assess the global range of motion for tRNAs. Biased simulations can be used to steer between known conformational stop points, while unbiased simulations allow for a general testing of conformational space previously unexplored. The unbiased molecular dynamics data describes the global conformational changes of tRNA on a sub-microsecond time scale for comparison with steered data. Additionally, the unbiased molecular dynamics data was used to identify putative contacts between tRNA and the ribosome during the accommodation step of translation. We found that the primary contact regions were H71 and H92 of the 50S subunit and ribosomal proteins L14 and L16.

No MeSH data available.