High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP.
Native data, anomalous data at three wavelengths and an independent peak-wavelength data set for SeMet-substituted protein have been collected from cryoprotected crystals of the TrpRS-adenylate product (TAM) complex to a resolution limit of 1.7 A. Independent phase sets were developed using SHARP an...
| Main Authors: | , , , , , , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2001
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| _version_ | 1797050894914682880 |
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| author | Retailleau, P Yin, Y Hu, M Roach, J Bricogne, G Vonrhein, C Roversi, P Blanc, E Sweet, R Carter, C |
| author_facet | Retailleau, P Yin, Y Hu, M Roach, J Bricogne, G Vonrhein, C Roversi, P Blanc, E Sweet, R Carter, C |
| author_sort | Retailleau, P |
| collection | OXFORD |
| description | Native data, anomalous data at three wavelengths and an independent peak-wavelength data set for SeMet-substituted protein have been collected from cryoprotected crystals of the TrpRS-adenylate product (TAM) complex to a resolution limit of 1.7 A. Independent phase sets were developed using SHARP and improved by solvent flipping with SOLOMON using molecular envelopes derived from experimental densities for, respectively, peak-wavelength SAD data from four different crystals, MAD data and their M(S)IRAS combinations with native data. Hendrickson-Lattman phase-probability coefficients from each phase set were used in BUSTER to drive maximum-likelihood refinements of well defined parts of the previously refined room-temperature 2.9 A structure. Maximum-entropy completion followed by manual rebuilding was then used to generate a model for the missing segments, bound ligand and solvent molecules. Surprisingly, peak-wavelength SAD experiments produced the smallest phase errors relative to the refined structures. Selenomethionylated models deviate from one another by 0.25 A and from the native model by 0.38 A, but all have r.m.s. deviations of approximately 1.0 A from the 2.9 A model. Difference Fourier calculations between amplitudes from the 300 K experiment and the new amplitudes at 100 K using 1.7 A model phases show no significant structural changes arising from temperature variation or addition of cryoprotectant. The main differences between low- and high-resolution structures arise from correcting side-chain rotamers in the core of the protein as well as on the surface. These changes improve various structure-validation criteria. |
| first_indexed | 2024-03-06T18:11:59Z |
| format | Journal article |
| id | oxford-uuid:0357749e-35cf-40f8-af19-35029b042f13 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T18:11:59Z |
| publishDate | 2001 |
| record_format | dspace |
| spelling | oxford-uuid:0357749e-35cf-40f8-af19-35029b042f132022-03-26T08:45:35ZHigh-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:0357749e-35cf-40f8-af19-35029b042f13EnglishSymplectic Elements at Oxford2001Retailleau, PYin, YHu, MRoach, JBricogne, GVonrhein, CRoversi, PBlanc, ESweet, RCarter, CNative data, anomalous data at three wavelengths and an independent peak-wavelength data set for SeMet-substituted protein have been collected from cryoprotected crystals of the TrpRS-adenylate product (TAM) complex to a resolution limit of 1.7 A. Independent phase sets were developed using SHARP and improved by solvent flipping with SOLOMON using molecular envelopes derived from experimental densities for, respectively, peak-wavelength SAD data from four different crystals, MAD data and their M(S)IRAS combinations with native data. Hendrickson-Lattman phase-probability coefficients from each phase set were used in BUSTER to drive maximum-likelihood refinements of well defined parts of the previously refined room-temperature 2.9 A structure. Maximum-entropy completion followed by manual rebuilding was then used to generate a model for the missing segments, bound ligand and solvent molecules. Surprisingly, peak-wavelength SAD experiments produced the smallest phase errors relative to the refined structures. Selenomethionylated models deviate from one another by 0.25 A and from the native model by 0.38 A, but all have r.m.s. deviations of approximately 1.0 A from the 2.9 A model. Difference Fourier calculations between amplitudes from the 300 K experiment and the new amplitudes at 100 K using 1.7 A model phases show no significant structural changes arising from temperature variation or addition of cryoprotectant. The main differences between low- and high-resolution structures arise from correcting side-chain rotamers in the core of the protein as well as on the surface. These changes improve various structure-validation criteria. |
| spellingShingle | Retailleau, P Yin, Y Hu, M Roach, J Bricogne, G Vonrhein, C Roversi, P Blanc, E Sweet, R Carter, C High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title | High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title_full | High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title_fullStr | High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title_full_unstemmed | High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title_short | High-resolution experimental phases for tryptophanyl-tRNA synthetase (TrpRS) complexed with tryptophanyl-5'AMP. |
| title_sort | high resolution experimental phases for tryptophanyl trna synthetase trprs complexed with tryptophanyl 5 amp |
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