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- PDB-8i70: Crystal structure of NADP-binding form of malonyl-CoA reductase C... -

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Basic information

Entry
Database: PDB / ID: 8i70
TitleCrystal structure of NADP-binding form of malonyl-CoA reductase C-domain from Chloroflexus aurantiacus
ComponentsShort-chain dehydrogenase/reductase SDR
KeywordsOXIDOREDUCTASE / NAD(P)-binding protein / Short-chain reductase
Function / homologyfatty acid elongation / oxidoreductase activity, acting on the CH-OH group of donors, NAD or NADP as acceptor / short chain dehydrogenase / Short-chain dehydrogenase/reductase SDR / NAD(P)-binding domain superfamily / NADP NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE / L(+)-TARTARIC ACID / Short-chain dehydrogenase/reductase SDR
Function and homology information
Biological speciesChloroflexus aurantiacus (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.9 Å
AuthorsKim, S. / Kim, K.-J.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Int J Biol Macromol / Year: 2023
Title: Cryo-EM structure of bifunctional malonyl-CoA reductase from Chloroflexus aurantiacus reveals a dynamic domain movement for high enzymatic activity.
Authors: Jae-Woo Ahn / Sangwoo Kim / Jiyeon Hong / Kyung-Jin Kim /
Abstract: The platform chemical 3-hydroxypropionic acid is used to synthesize various valuable materials, including bioplastics. Bifunctional malonyl-CoA reductase is a key enzyme in 3-hydroxypropionic acid ...The platform chemical 3-hydroxypropionic acid is used to synthesize various valuable materials, including bioplastics. Bifunctional malonyl-CoA reductase is a key enzyme in 3-hydroxypropionic acid biosynthesis as it catalyzes the two-step reduction of malonyl-CoA to malonate semialdehyde to 3-hydroxypropionic acid. Here, we report the cryo-EM structure of a full-length malonyl-CoA reductase protein from Chloroflexus aurantiacus (CaMCR). The EM model of CaMCR reveals a tandem helix architecture comprising an N-terminal (CaMCR) and a C-terminal (CaMCR) domain. The CaMCR model also revealed that the enzyme undergoes a dynamic domain movement between CaMCR and CaMCR due to the presence of a flexible linker between these two domains. Increasing the flexibility and extension of the linker resulted in a twofold increase in enzyme activity, indicating that for CaMCR, domain movement is crucial for high enzyme activity. We also describe the structural features of CaMCR and CaMCR. This study reveals the protein structures underlying the molecular mechanism of CaMCR and thereby provides valuable information for future enzyme engineering to improve the productivity of 3-hydroxypropionic acid.
History
DepositionJan 30, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 14, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Short-chain dehydrogenase/reductase SDR
hetero molecules


Theoretical massNumber of molelcules
Total (without water)76,6406
Polymers75,3551
Non-polymers1,2865
Water8,647480
1
A: Short-chain dehydrogenase/reductase SDR
hetero molecules

A: Short-chain dehydrogenase/reductase SDR
hetero molecules


Theoretical massNumber of molelcules
Total (without water)153,28112
Polymers150,7092
Non-polymers2,57210
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_656-x+1,y,-z+11
Buried area10710 Å2
ΔGint-24 kcal/mol
Surface area45690 Å2
MethodPISA
Unit cell
Length a, b, c (Å)85.675, 139.644, 73.565
Angle α, β, γ (deg.)90.00, 97.71, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Protein Short-chain dehydrogenase/reductase SDR / Malonyl-CoA reductase


Mass: 75354.688 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chloroflexus aurantiacus (bacteria) / Gene: Caur_2614 / Plasmid: pET28 / Production host: Escherichia coli BL21(DE3) (bacteria)
References: UniProt: A9WIU3, malonyl-CoA reductase (malonate semialdehyde-forming)
#2: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL / Glycerol


Mass: 92.094 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C3H8O3
#3: Chemical ChemComp-TLA / L(+)-TARTARIC ACID / Tartaric acid


Mass: 150.087 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C4H6O6 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-NAP / NADP NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE / 2'-MONOPHOSPHOADENOSINE 5'-DIPHOSPHORIBOSE / Nicotinamide adenine dinucleotide phosphate


Mass: 743.405 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C21H28N7O17P3
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 480 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.02 Å3/Da / Density % sol: 59.36 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, hanging drop / pH: 7 / Details: 0.1 M ammonium tartrate, 0.1 M Tris-Cl

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: PAL/PLS / Beamline: 7A (6B, 6C1) / Wavelength: 0.97934 Å
DetectorType: ADSC QUANTUM 270 / Detector: CCD / Date: Nov 18, 2016
RadiationMonochromator: DCM Si (111) Crystal / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97934 Å / Relative weight: 1
ReflectionResolution: 1.9→50 Å / Num. obs: 67574 / % possible obs: 98.8 % / Redundancy: 2.1 % / Rmerge(I) obs: 0.064 / Χ2: 0.046 / Net I/σ(I): 12.9 / Num. measured all: 279616
Reflection shell
Resolution (Å)Redundancy (%)Rmerge(I) obsNum. unique obsΧ2Diffraction-ID% possible all
1.9-1.932.10.23264831.573198.1
1.93-1.972.10.19665491.561198.3
1.97-2.012.10.16665821.585198.5
2.01-2.052.10.14565961.616198.5
2.05-2.092.10.1365961.564198.5
2.09-2.142.10.10965131.581198.7
2.14-2.192.10.09266391.571198.7
2.19-2.252.10.08765731.632198.8
2.25-2.322.10.07765801.611199
2.32-2.392.10.07166281.62199
2.39-2.482.10.06466561.575199.1
2.48-2.582.10.05865561.6199.1
2.58-2.72.10.05566131.69199.3
2.7-2.842.10.04866911.646199.4
2.84-3.022.10.04366361.585199.6
3.02-3.252.10.0466451.635199.6
3.25-3.582.10.03766761.796199.7
3.58-4.092.10.03666651.948199.7
4.09-5.162.10.03566031.921199.3
5.16-5020.03963792.398195.5

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Processing

Software
NameVersionClassification
REFMACv5.8.0267refinement
HKL-2000data scaling
HKL-2000data reduction
MOLREPphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 8I6Z

8i6z


Resolution: 1.9→26.54 Å / Cor.coef. Fo:Fc: 0.969 / Cor.coef. Fo:Fc free: 0.951 / SU B: 2.322 / SU ML: 0.069 / Cross valid method: THROUGHOUT / ESU R: 0.109 / ESU R Free: 0.11 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.19129 3556 5.3 %RANDOM
Rwork0.15229 ---
obs0.15425 63699 99.51 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 26.733 Å2
Baniso -1Baniso -2Baniso -3
1-1.34 Å2-0 Å2-0.86 Å2
2---0.57 Å20 Å2
3----0.51 Å2
Refinement stepCycle: 1 / Resolution: 1.9→26.54 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5080 0 84 480 5644
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0130.0135259
X-RAY DIFFRACTIONr_bond_other_d0.0010.0145034
X-RAY DIFFRACTIONr_angle_refined_deg1.7281.6577143
X-RAY DIFFRACTIONr_angle_other_deg1.4761.57411537
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.225657
X-RAY DIFFRACTIONr_dihedral_angle_2_deg30.78720.068296
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.33415863
X-RAY DIFFRACTIONr_dihedral_angle_4_deg13.661559
X-RAY DIFFRACTIONr_chiral_restr0.0870.2699
X-RAY DIFFRACTIONr_gen_planes_refined0.010.025978
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021227
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it2.4772.4872631
X-RAY DIFFRACTIONr_mcbond_other2.4762.4872630
X-RAY DIFFRACTIONr_mcangle_it3.1863.7153287
X-RAY DIFFRACTIONr_mcangle_other3.1863.7153288
X-RAY DIFFRACTIONr_scbond_it3.9643.0192628
X-RAY DIFFRACTIONr_scbond_other3.9643.0192626
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other5.9344.3373857
X-RAY DIFFRACTIONr_long_range_B_refined7.41631.3765928
X-RAY DIFFRACTIONr_long_range_B_other7.34431.0755850
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.9→1.946 Å
RfactorNum. reflection% reflection
Rfree0.252 265 -
Rwork0.188 4545 -
obs--96.2 %

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