+
Open data
-
Basic information
Entry | Database: PDB / ID: 6kpi | ||||||
---|---|---|---|---|---|---|---|
Title | 298 K cryoEM structure of Sso-KARI in complex with Mg2+ | ||||||
![]() | Ketol-acid reductoisomerase | ||||||
![]() | ISOMERASE / Complex | ||||||
Function / homology | ![]() ketol-acid reductoisomerase (NADP+) / ketol-acid reductoisomerase activity / valine biosynthetic process / isoleucine biosynthetic process / isomerase activity / metal ion binding / cytosol Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.43 Å | ||||||
![]() | Chen, C.Y. / Chang, Y.C. / Lin, B.L. / Huang, C.H. / Tsai, M.D. | ||||||
![]() | ![]() Title: Temperature-Resolved Cryo-EM Uncovers Structural Bases of Temperature-Dependent Enzyme Functions. Authors: Chin-Yu Chen / Yuan-Chih Chang / Bo-Lin Lin / Chun-Hsiang Huang / Ming-Daw Tsai / ![]() Abstract: Protein functions are temperature-dependent, but protein structures are usually solved at a single (often low) temperature because of limitations on the conditions of crystal growth or protein ...Protein functions are temperature-dependent, but protein structures are usually solved at a single (often low) temperature because of limitations on the conditions of crystal growth or protein vitrification. Here we demonstrate the feasibility of solving cryo-EM structures of proteins vitrified at high temperatures, solve 12 structures of an archaeal ketol-acid reductoisomerase (KARI) vitrified at 4-70 °C, and show that structures of both the Mg form (KARI:2Mg) and its ternary complex (KARI:2Mg:NADH:inhibitor) are temperature-dependent in correlation with the temperature dependence of enzyme activity. Furthermore, structural analyses led to dissection of the induced-fit mechanism into ligand-induced and temperature-induced effects and to capture of temperature-resolved intermediates of the temperature-induced conformational change. The results also suggest that it is preferable to solve cryo-EM structures of protein complexes at functional temperatures. These studies should greatly expand the landscapes of protein structure-function relationships and enhance the mechanistic analysis of enzymatic functions. | ||||||
History |
|
-
Structure visualization
Movie |
![]() |
---|---|
Structure viewer | Molecule: ![]() ![]() |
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 1.2 MB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | 1 MB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
---|
-Related structure data
Related structure data | ![]() 0747MC ![]() 0740C ![]() 0742C ![]() 0743C ![]() 0746C ![]() 0748C ![]() 0749C ![]() 0750C ![]() 0751C ![]() 0752C ![]() 0753C ![]() 0754C ![]() 6kouC ![]() 6kpaC ![]() 6kpeC ![]() 6kphC ![]() 6kpjC ![]() 6kpkC ![]() 6kq4C ![]() 6kq8C ![]() 6kqjC ![]() 6kqkC ![]() 6kqoC C: citing same article ( M: map data used to model this data |
---|---|
Similar structure data |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 |
|
-
Components
#1: Protein | Mass: 37229.855 Da / Num. of mol.: 12 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() #2: Chemical | ChemComp-MG / Has ligand of interest | Y | |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-
Sample preparation
Component | Name: KARI-Mg2+ complex / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
---|---|
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 / Details: 20 mM Tris-Cl, pH 7.5, 50 mM NaCl and 5 mM MgCl2 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 52 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k) |
-
Processing
CTF correction | Type: PHASE FLIPPING ONLY |
---|---|
3D reconstruction | Resolution: 2.43 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 81592 / Symmetry type: POINT |