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- PDB-6jd2: Crystal structure of Sulfolobus solfataricus ketol-acid reductois... -
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Open data
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Basic information
Entry | Database: PDB / ID: 6jd2 | |||||||||
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Title | Crystal structure of Sulfolobus solfataricus ketol-acid reductoisomerase (Sso-KARI) in complex with Mg2+ at pH8.5 | |||||||||
![]() | Putative ketol-acid reductoisomerase 2 | |||||||||
![]() | ISOMERASE / Bi-specific / Thermostable / Reductoisomerase / Magnesium-dependent / Dodecamer / Knotted protein | |||||||||
Function / homology | ![]() ketol-acid reductoisomerase (NADP+) / ketol-acid reductoisomerase activity / valine biosynthetic process / isoleucine biosynthetic process / amino acid biosynthetic process / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ![]() ![]() ![]() | |||||||||
![]() | Chen, C.Y. / Chang, Y.C. / Lin, K.F. / Huang, C.H. / Lin, B.L. / Ko, T.P. / Hsieh, D.L. / Tsai, M.D. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Use of Cryo-EM To Uncover Structural Bases of pH Effect and Cofactor Bispecificity of Ketol-Acid Reductoisomerase. Authors: Chin-Yu Chen / Yuan-Chih Chang / Bo-Lin Lin / Kuan-Fu Lin / Chun-Hsiang Huang / Dong-Lin Hsieh / Tzu-Ping Ko / Ming-Daw Tsai / ![]() Abstract: While cryo-EM is revolutionizing structural biology, its impact on enzymology is yet to be fully demonstrated. The ketol-acid reductoisomerase (KARI) catalyzes conversion of (2 S)-acetolactate or (2 ...While cryo-EM is revolutionizing structural biology, its impact on enzymology is yet to be fully demonstrated. The ketol-acid reductoisomerase (KARI) catalyzes conversion of (2 S)-acetolactate or (2 S)-aceto-2-hydroxybutyrate to 2,3-dihydroxy-3-alkylbutyrate. We found that KARI from archaea Sulfolobus solfataricus (Sso-KARI) is unusual in being a dodecamer, bispecific to NADH and NADPH, and losing activity above pH 7.8. While crystals were obtainable only at pH 8.5, cryo-EM structures were solved at pH 7.5 and 8.5 for Sso-KARI:2Mg. The results showed that the distances of the two catalytic Mg ions are lengthened in both structures at pH 8.5. We next solved cryo-EM structures of two Sso-KARI complexes, with NADH+inhibitor and NADPH+inhibitor at pH 7.5, which indicate that the bispecificity can be attributed to a unique asparagine at the cofactor binding loop. Unexpectedly, Sso-KARI also differs from other KARI enzymes in lacking "induced-fit", reflecting structural rigidity. Thus, cryo-EM is powerful for structural and mechanistic enzymology. | |||||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 767.1 KB | Display | ![]() |
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PDB format | ![]() | 636.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 545.2 KB | Display | ![]() |
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Full document | ![]() | 598.2 KB | Display | |
Data in XML | ![]() | 139.1 KB | Display | |
Data in CIF | ![]() | 191.9 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 9798C ![]() 9799C ![]() 9800C ![]() 9801C ![]() 6jcvC ![]() 6jcwC ![]() 6jczC ![]() 6jd1C ![]() 1np3S S: Starting model for refinement C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Unit cell |
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Components
#1: Protein | Mass: 37229.855 Da / Num. of mol.: 12 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Strain: ATCC 35092 / DSM 1617 / JCM 11322 / P2 / Gene: ilvC2, ilvC-2, SSO1322 / Production host: ![]() ![]() References: UniProt: Q97YJ9, ketol-acid reductoisomerase (NADP+) #2: Chemical | ChemComp-BME / #3: Chemical | ChemComp-MG / #4: Water | ChemComp-HOH / | |
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-Experimental details
-Experiment
Experiment | Method: ![]() |
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Sample preparation
Crystal | Density Matthews: 2.83 Å3/Da / Density % sol: 56.53 % |
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Crystal grow | Temperature: 298 K / Method: vapor diffusion, sitting drop / pH: 8.5 Details: 15% PEG5000MME, 0.15M KSCN, 0.1M Tis-Cl, pH8.5 Cryo-protectant: 20% ethylene glycol |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: ![]() ![]() ![]() |
Detector | Type: ADSC QUANTUM 315r / Detector: CCD / Date: Oct 25, 2014 |
Radiation | Monochromator: Horizontally Focusing Single Crystal Monochromator Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.97622 Å / Relative weight: 1 |
Reflection | Resolution: 2.53→30 Å / Num. obs: 165970 / % possible obs: 99.4 % / Redundancy: 4.1 % / Rmerge(I) obs: 0.069 / Net I/σ(I): 22.2 |
Reflection shell | Resolution: 2.52→2.61 Å / Redundancy: 4.1 % / Rmerge(I) obs: 0.582 / Num. unique obs: 16118 / % possible all: 97.5 |
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Processing
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Refinement | Method to determine structure: ![]() Starting model: 1NP3 Resolution: 2.53→30 Å / Cross valid method: FREE R-VALUE
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Refinement step | Cycle: LAST / Resolution: 2.53→30 Å
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LS refinement shell | Resolution: 2.53→2.61 Å
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