PDB-8w7e: Design, synthesis and biological evaluations of novel small molec...

Basic information

• Entry: Database: PDB / ID: 8w7e
• Title: Design, synthesis and biological evaluations of novel small molecular hyper-activators of human caseinolytic peptidase P (hClpP)
• Components: ATP-dependent Clp protease proteolytic subunit, mitochondrial
• Keywords: APOPTOSIS / F20 / activators / hClpP

Function / homology

Function:

membrane protein proteolysis / mitochondrial protein catabolic process / endopeptidase Clp / endopeptidase Clp complex / ATP-dependent peptidase activity / protein quality control for misfolded or incompletely synthesized proteins / Mitochondrial protein degradation / proteolysis involved in protein catabolic process / peptidase activity / ATPase binding / endopeptidase activity / mitochondrial matrix / serine-type endopeptidase activity / mitochondrion / proteolysis / identical protein binding

Domain/homology:

ClpP, Ser active site / Endopeptidase Clp serine active site. / ClpP, histidine active site / Endopeptidase Clp histidine active site. / ATP-dependent Clp protease proteolytic subunit / Clp protease proteolytic subunit /Translocation-enhancing protein TepA / Clp protease / ClpP/crotonase-like domain superfamily

Component:

: / ATP-dependent Clp protease proteolytic subunit, mitochondrial

• Biological species: Homo sapiens (human)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.8 Å
• Authors: Jiang, J.-X. / Ding, H. / Chen, M.-R. / Lu, M.-L. / Sun, H.-Y. / Xiao, Y.-B.

Funding support

1items

Organization	Grant number	Country
Not funded

• Citation: Journal: To Be Published; Title: Design, synthesis and biological evaluations of novel small molecular hyper-activators of human caseinolytic peptidase P (hClpP); Authors: Jiang, J.-X. / Ding, H. / Chen, M.-R. / Lu, M.-L. / Sun, H.-Y. / Xiao, Y.-B.

History

Deposition	Aug 30, 2023	Deposition site: PDBJ / Processing site: PDBC
Revision 1.0	Sep 4, 2024	Provider: repository / Type: Initial release

Assembly

Deposited unit

A: ATP-dependent Clp protease proteolytic subunit, mitochondrial

B: ATP-dependent Clp protease proteolytic subunit, mitochondrial

C: ATP-dependent Clp protease proteolytic subunit, mitochondrial

D: ATP-dependent Clp protease proteolytic subunit, mitochondrial

E: ATP-dependent Clp protease proteolytic subunit, mitochondrial

F: ATP-dependent Clp protease proteolytic subunit, mitochondrial

G: ATP-dependent Clp protease proteolytic subunit, mitochondrial

H: ATP-dependent Clp protease proteolytic subunit, mitochondrial

I: ATP-dependent Clp protease proteolytic subunit, mitochondrial

J: ATP-dependent Clp protease proteolytic subunit, mitochondrial

K: ATP-dependent Clp protease proteolytic subunit, mitochondrial

L: ATP-dependent Clp protease proteolytic subunit, mitochondrial

M: ATP-dependent Clp protease proteolytic subunit, mitochondrial

N: ATP-dependent Clp protease proteolytic subunit, mitochondrial

hetero molecules

Summary:

• 347 kDa, 14 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	346,841	28
Polymers	340,536	14
Non-polymers	6,305	14
Water	0	0

1

A: ATP-dependent Clp protease proteolytic subunit, mitochondrial

B: ATP-dependent Clp protease proteolytic subunit, mitochondrial

C: ATP-dependent Clp protease proteolytic subunit, mitochondrial

D: ATP-dependent Clp protease proteolytic subunit, mitochondrial

F: ATP-dependent Clp protease proteolytic subunit, mitochondrial

G: ATP-dependent Clp protease proteolytic subunit, mitochondrial

H: ATP-dependent Clp protease proteolytic subunit, mitochondrial

I: ATP-dependent Clp protease proteolytic subunit, mitochondrial

K: ATP-dependent Clp protease proteolytic subunit, mitochondrial

M: ATP-dependent Clp protease proteolytic subunit, mitochondrial

N: ATP-dependent Clp protease proteolytic subunit, mitochondrial

hetero molecules

J: ATP-dependent Clp protease proteolytic subunit, mitochondrial

L: ATP-dependent Clp protease proteolytic subunit, mitochondrial

hetero molecules

E: ATP-dependent Clp protease proteolytic subunit, mitochondrial

hetero molecules

Summary:

• defined by author&software
• Evidence: gel filtration
• 347 kDa, 14 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	346,841	28
Polymers	340,536	14
Non-polymers	6,305	14
Water	0

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	3_445	x-1/2,y-1/2,z	1
crystal symmetry operation	4_445	-x-1/2,y-1/2,-z	1

Calculated values:

Buried area	48380 Å2
ΔGint	-240 kcal/mol
Surface area	84330 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	164.767, 119.217, 152.807
Angle α, β, γ (deg.)	90.00, 100.82, 90.00
Int Tables number	5
Space group name H-M	C121

Components

#1: Protein

A B C D E F G H I J K L M N
ATP-dependent Clp protease proteolytic subunit, mitochondrial / Endopeptidase Clp

Details:

Mass: 24324.004 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Details: Author stated: The first two amino acid (Gly and Ser) is a linker proteased by SUMO protease. The third amino acid (N0.57 Ser) belongs to disorder region.
Source: (gene. exp.) Homo sapiens (human) / Gene: CLPP / Production host: Escherichia coli (E. coli) / References: UniProt: Q16740, endopeptidase Clp

#2: Chemical

A-301 B-301 C-301 D-301 E-301 E-302 F-301 G-301 H-301 I-301 K-301 L-301 M-301 N-301
ChemComp-9I3 / 3-[(3-chlorophenyl)methyl]-6-[(4-chlorophenyl)methyl]-2,4-dihydro-1H-pyrido[2,3-c][2,7]naphthyridin-5-one

Details:

Mass: 450.360 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: C₂₅H₂₁Cl₂N₃O

• Has ligand of interest: N

Experimental details

Experiment

• Experiment: Method: X-RAY DIFFRACTION / Number of used crystals: 1

Sample preparation

• Crystal: Density Matthews: 2.21 ų/Da / Density % sol: 44.3 %
• Crystal grow: Temperature: 289 K / Method: vapor diffusion, hanging drop / pH: 6 ; Details: 6% (w/v) PEG 3350, 200 mM KCl, and 100 mM NaAc (pH 6.0)

Data collection

• Diffraction: Mean temperature: 100 K / Serial crystal experiment: N
• Diffraction source: Source: SYNCHROTRON / Site: SSRF / Beamline: BL19U1 / Wavelength: 1 Å
• Detector: Type: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jul 3, 2021
• Radiation: Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 1 Å / Relative weight: 1
• Reflection: Resolution: 2.8→46.65 Å / Num. obs: 140112 / % possible obs: 99.62 % / Redundancy: 3.7 % / CC_1/2: 0.994 / CC star: 0.998 / R_pim(I) all: 0.0842 / Net I/σ(I): 8.97
• Reflection shell: Resolution: 2.8→2.9 Å / Num. unique obs: 7203 / CC_1/2: 0.566 / CC star: 0.85 / R_pim(I) all: 0.6652

Processing

Software

Name	Version	Classification
PHENIX	(1.18.2_3874: ???)	refinement
HKL-3000		data reduction
HKL-3000		data scaling
PHENIX		phasing

Refinement

Method to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.8→46.65 Å / SU ML: 0.46 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 30.4 / Stereochemistry target values: ML

	Rfactor	Num. reflection	% reflection
Rfree	0.2738	3841	2.74 %
Rwork	0.2661	-	-
obs	0.2663	140112	99.63 %

• Solvent computation: Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL

Refinement step

Cycle: LAST / Resolution: 2.8→46.65 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	18610	0	434	0	19044

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.006	19477
X-RAY DIFFRACTION	f_angle_d	0.889	26423
X-RAY DIFFRACTION	f_dihedral_angle_d	17.455	7217
X-RAY DIFFRACTION	f_chiral_restr	0.052	3035
X-RAY DIFFRACTION	f_plane_restr	0.005	3277

LS refinement shell

Resolution (Å)	Rfactor Rfree	Num. reflection Rfree	Rfactor Rwork	Num. reflection Rwork	Refine-ID	% reflection obs (%)
2.8-2.84	0.3254	139	0.3711	5082	X-RAY DIFFRACTION	100
2.84-2.87	0.4044	143	0.3704	4988	X-RAY DIFFRACTION	100
2.87-2.91	0.3498	146	0.3677	5079	X-RAY DIFFRACTION	100
2.91-2.95	0.3307	140	0.3413	5037	X-RAY DIFFRACTION	100
2.95-3	0.3292	145	0.3454	5117	X-RAY DIFFRACTION	100
3-3.04	0.3443	142	0.3296	5037	X-RAY DIFFRACTION	100
3.04-3.09	0.3527	145	0.3412	5055	X-RAY DIFFRACTION	100
3.09-3.15	0.3179	142	0.3322	5052	X-RAY DIFFRACTION	100
3.15-3.21	0.3778	140	0.3402	5065	X-RAY DIFFRACTION	100
3.21-3.27	0.3732	136	0.3571	5007	X-RAY DIFFRACTION	98
3.27-3.33	0.3131	143	0.3109	5013	X-RAY DIFFRACTION	100
3.33-3.41	0.323	144	0.3128	5140	X-RAY DIFFRACTION	100
3.41-3.49	0.3242	138	0.3056	5014	X-RAY DIFFRACTION	100
3.49-3.57	0.2827	139	0.2971	5020	X-RAY DIFFRACTION	100
3.57-3.67	0.3261	142	0.2989	5031	X-RAY DIFFRACTION	100
3.67-3.78	0.3269	148	0.2841	5129	X-RAY DIFFRACTION	100
3.78-3.9	0.2887	138	0.2785	4990	X-RAY DIFFRACTION	100
3.9-4.04	0.2906	143	0.2756	5089	X-RAY DIFFRACTION	100
4.04-4.2	0.2799	141	0.2548	5028	X-RAY DIFFRACTION	100
4.2-4.39	0.2719	145	0.2374	5065	X-RAY DIFFRACTION	100
4.39-4.62	0.2612	149	0.2401	5074	X-RAY DIFFRACTION	99
4.62-4.91	0.2422	137	0.226	4968	X-RAY DIFFRACTION	99
4.91-5.29	0.2309	139	0.2458	5049	X-RAY DIFFRACTION	99
5.29-5.82	0.2512	145	0.2448	5030	X-RAY DIFFRACTION	100
5.82-6.66	0.2551	144	0.2391	5057	X-RAY DIFFRACTION	100
6.66-8.38	0.2193	144	0.2092	5055	X-RAY DIFFRACTION	100
8.39-46.65	0.167	144	0.1764	5000	X-RAY DIFFRACTION	99

Refinement TLS params.

Method: refined / Origin x: -33.2964 Å / Origin y: 35.5922 Å / Origin z: 37.0644 Å

	11	12	13	21	22	23	31	32	33
T	0.3407 Å2	-0.0714 Å2	0.0101 Å2	-	0.3966 Å2	-0.022 Å2	-	-	0.4548 Å2
L	0.1146 °2	0.0193 °2	0.0627 °2	-	0.1249 °2	-0.0726 °2	-	-	0.487 °2
S	0.049 Å °	-0.0642 Å °	0.0211 Å °	0.0113 Å °	-0.0593 Å °	-0.0056 Å °	0.0326 Å °	0.014 Å °	0.0121 Å °

• Refinement TLS group: Selection details: all