PDB-7mdb: Crystal structure of Staphylococcus aureus cystathionine gamma ly...

Basic information

• Entry: Database: PDB / ID: 7mdb
• Title: Crystal structure of Staphylococcus aureus cystathionine gamma lyase holoenzyme Y103A mutant co-crystallized with NL2
• Components: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase
• Keywords: LYASE / Amino-acid biosynthesis / Cysteine biosynthesis / Hydrogen sulfide production / PLP dependent enzyme

Function / homology

Function:

cystathionine beta-lyase / cystathionine gamma-synthase / cystathionine gamma-lyase / cystathionine gamma-synthase activity / cystathionine gamma-lyase activity / cysteine biosynthetic process via cystathionine / transsulfuration / pyridoxal phosphate binding / identical protein binding / cytoplasm

Domain/homology:

: / Cys/Met metabolism enzymes pyridoxal-phosphate attachment site. / Cys/Met metabolism, pyridoxal phosphate-dependent enzyme / Cys/Met metabolism PLP-dependent enzyme / Pyridoxal phosphate-dependent transferase, small domain / Pyridoxal phosphate-dependent transferase, major domain / Pyridoxal phosphate-dependent transferase

Component:

CACODYLATE ION / PYRIDOXAL-5'-PHOSPHATE / Chem-YXA / Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

• Biological species: Staphylococcus aureus (bacteria)
• Method: X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.8 Å
• Authors: Nuthanakanti, A. / Serganov, A. / Kaushik, A.

Funding support

United States, 3items

Organization	Grant number	Country
Department of Defense (DOD, United States)	PR171734	United States
Department of Defense (DOD, United States)	PR171734P1	United States
Howard Hughes Medical Institute (HHMI)		United States

• Citation: Journal: Science / Year: 2021; Title: Inhibitors of bacterial H 2 S biogenesis targeting antibiotic resistance and tolerance.; Authors: Shatalin, K. / Nuthanakanti, A. / Kaushik, A. / Shishov, D. / Peselis, A. / Shamovsky, I. / Pani, B. / Lechpammer, M. / Vasilyev, N. / Shatalina, E. / Rebatchouk, D. / Mironov, A. / Fedichev, P. / Serganov, A. / Nudler, E.

History

Deposition	Apr 3, 2021	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Jun 23, 2021	Provider: repository / Type: Initial release
Revision 1.1	Oct 18, 2023	Group: Data collection / Database references / Refinement description Category: chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.2	Nov 15, 2023	Group: Data collection / Category: chem_comp_atom / chem_comp_bond / Item: _chem_comp_atom.atom_id / _chem_comp_bond.atom_id_2

Assembly

Deposited unit

H: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

hetero molecules

Summary:

• 42.8 kDa, 1 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	42,837	10
Polymers	41,418	1
Non-polymers	1,420	9
Water	6,341	352

1

H: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

hetero molecules

H: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

hetero molecules

H: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

hetero molecules

H: Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase

hetero molecules

Summary:

• defined by author&software
• Evidence: gel filtration
• 171 kDa, 4 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	171,349	40
Polymers	165,670	4
Non-polymers	5,678	36
Water	72	4

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1
crystal symmetry operation	8_665	-y+1,-x+1,-z	1
crystal symmetry operation	10_665	-x+1,-y+1,z	1
crystal symmetry operation	15_555	y,x,-z	1

Calculated values:

Buried area	27450 Å2
ΔGint	-129 kcal/mol
Surface area	43960 Å2
Method	PISA

Unit cell

Length a, b, c (Å)	105.119, 105.119, 287.790
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	98
Space group name H-M	I4122
Space group name Hall	I4bw2bw
Symmetry operation	#1: x,y,z #2: -y+1/2,x,z+3/4 #3: y+1/2,-x,z+3/4 #4: x+1/2,-y,-z+3/4 #5: -x+1/2,y,-z+3/4 #6: -x,-y,z #7: y,x,-z #8: -y,-x,-z #9: x+1/2,y+1/2,z+1/2 #10: -y+1,x+1/2,z+5/4 #11: y+1,-x+1/2,z+5/4 #12: x+1,-y+1/2,-z+5/4 #13: -x+1,y+1/2,-z+5/4 #14: -x+1/2,-y+1/2,z+1/2 #15: y+1/2,x+1/2,-z+1/2 #16: -y+1/2,-x+1/2,-z+1/2

Components on special symmetry positions

ID	Model	Components
1	1	H-729- HOH
2	1	H-833- HOH
3	1	H-850- HOH

Components

Protein , 1 types, 1 molecules H

#1: Protein

H Bifunctional cystathionine gamma-lyase/homocysteine desulfhydrase / Cystathionine beta-lyase / Cystathionine gamma-lyase / Cystathionine gamma-synthase

Details:

Mass: 41417.617 Da / Num. of mol.: 1 / Mutation: Y103A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Staphylococcus aureus (bacteria) / Gene: mccB / Production host: Escherichia coli BL21(DE3) (bacteria)
References: UniProt: X5E0F1, cystathionine gamma-lyase, cystathionine beta-lyase, cystathionine gamma-synthase

Non-polymers , 6 types, 361 molecules

#2: Chemical

H-401 H-402 ChemComp-YXA / 5-[(6-bromo-1H-indol-1-yl)methyl]-2-methylfuran-3-carboxylic acid

Details:

Mass: 334.165 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₁₅H₁₂BrNO₃ / Feature type: SUBJECT OF INVESTIGATION

#3: Chemical

H-403 ChemComp-PLP / PYRIDOXAL-5'-PHOSPHATE / VITAMIN B6 Phosphate

Details:

Mass: 247.142 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₈H₁₀NO₆P

#4: Chemical

H-404 H-405 ChemComp-CAC / CACODYLATE ION / dimethylarsinate

Details:

Mass: 136.989 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₂H₆AsO₂

#5: Chemical

H-406 H-407 ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL

Details:

Mass: 92.094 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C₃H₈O₃

#6: Chemical

H-408 H-409 ChemComp-NA / SODIUM ION

Details:

Mass: 22.990 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Na

#7: Water

ChemComp-HOH / water

Details:

Mass: 18.015 Da / Num. of mol.: 352 / Source method: isolated from a natural source / Formula: H₂O

Details

• Has ligand of interest: Y

Experimental details

Experiment

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

Sample preparation

• Crystal: Density Matthews: 4.76 ų/Da / Density % sol: 74.18 %
• Crystal grow: Temperature: 291 K / Method: vapor diffusion, hanging drop / pH: 6.5 ; Details: 0.1 M sodium cacodylate , pH 6.5, 1.3 M tri-sodium citrate

Data collection

• Diffraction: Mean temperature: 100 K / Serial crystal experiment: N
• Diffraction source: Source: SYNCHROTRON / Site: NSLS-II / Beamline: 17-ID-2 / Wavelength: 0.91956 Å
• Detector: Type: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Jan 31, 2020
• Radiation: Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
• Radiation wavelength: Wavelength: 0.91956 Å / Relative weight: 1
• Reflection: Resolution: 1.8→30 Å / Num. obs: 74732 / % possible obs: 100 % / Redundancy: 8.9 % / B_iso Wilson estimate: 27.83 Ų / CC_1/2: 0.989 / R_merge(I) obs: 0.104 / R_pim(I) all: 0.037 / Net I/σ(I): 38.6
• Reflection shell: Resolution: 1.8→1.83 Å / Redundancy: 7.3 % / R_merge(I) obs: 1.409 / Mean I/σ(I) obs: 1.6 / Num. unique obs: 3667 / CC_1/2: 0.748 / R_pim(I) all: 0.539 / % possible all: 99.9

Processing

Software

Name	Version	Classification
PHENIX	1.14_3260	refinement
PHENIX	1.14_3260	refinement
autoPROC	1.14_3260	data processing
HKL-2000		data reduction
HKL-2000		data scaling
PHASER		phasing

Refinement

Method to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entry 4IXZ

4ixz

Resolution: 1.8→29.69 Å / SU ML: 0.232 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 20.5101 / Stereochemistry target values: CDL v1.2

	Rfactor	Num. reflection	% reflection
Rfree	0.1894	3801	2.67 %
Rwork	0.1697	138407	-
obs	0.1702	74627	99.6 %

• Solvent computation: Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
• Displacement parameters: B_iso mean: 32.31 Ų

Refinement step

Cycle: LAST / Resolution: 1.8→29.69 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	2899	0	79	352	3330

Refine LS restraints

Refine-ID	Type	Dev ideal	Number
X-RAY DIFFRACTION	f_bond_d	0.0071	3086
X-RAY DIFFRACTION	f_angle_d	1.0109	4214
X-RAY DIFFRACTION	f_chiral_restr	0.0591	496
X-RAY DIFFRACTION	f_plane_restr	0.0058	535
X-RAY DIFFRACTION	f_dihedral_angle_d	17.3311	1106

LS refinement shell

Resolution (Å)	Rfactor Rfree	Num. reflection Rfree	Rfactor Rwork	Num. reflection Rwork	Refine-ID	% reflection obs (%)
1.8-1.82	0.4521	129	0.3887	4822	X-RAY DIFFRACTION	94.04
1.82-1.85	0.341	146	0.3569	5160	X-RAY DIFFRACTION	99.59
1.85-1.87	0.3117	143	0.3209	5066	X-RAY DIFFRACTION	99.69
1.87-1.9	0.3004	138	0.2883	5129	X-RAY DIFFRACTION	99.58
1.9-1.93	0.2534	145	0.2747	5182	X-RAY DIFFRACTION	99.91
1.93-1.96	0.2948	145	0.2553	5103	X-RAY DIFFRACTION	99.89
1.96-1.99	0.2568	139	0.2356	5171	X-RAY DIFFRACTION	99.89
1.99-2.02	0.2142	141	0.2175	5132	X-RAY DIFFRACTION	99.62
2.02-2.06	0.2247	140	0.2189	5124	X-RAY DIFFRACTION	99.83
2.06-2.1	0.2236	143	0.2131	5142	X-RAY DIFFRACTION	99.87
2.1-2.14	0.2315	142	0.1928	5148	X-RAY DIFFRACTION	99.94
2.14-2.19	0.2381	142	0.1871	5166	X-RAY DIFFRACTION	99.83
2.19-2.24	0.2197	140	0.184	5109	X-RAY DIFFRACTION	99.81
2.24-2.3	0.1956	138	0.1839	5089	X-RAY DIFFRACTION	99.81
2.3-2.36	0.2004	141	0.1801	5180	X-RAY DIFFRACTION	99.94
2.36-2.43	0.2252	143	0.1821	5132	X-RAY DIFFRACTION	99.94
2.43-2.51	0.2052	140	0.176	5156	X-RAY DIFFRACTION	99.91
2.51-2.6	0.2122	138	0.1777	5131	X-RAY DIFFRACTION	99.91
2.6-2.7	0.2002	145	0.1702	5161	X-RAY DIFFRACTION	100
2.7-2.82	0.1658	144	0.1716	5136	X-RAY DIFFRACTION	99.91
2.82-2.97	0.1986	140	0.1711	5153	X-RAY DIFFRACTION	99.96
2.97-3.16	0.2044	142	0.1641	5110	X-RAY DIFFRACTION	99.87
3.16-3.4	0.1619	140	0.1486	5157	X-RAY DIFFRACTION	99.79
3.4-3.74	0.1431	141	0.1354	5128	X-RAY DIFFRACTION	99.81
3.74-4.28	0.1109	142	0.1188	5145	X-RAY DIFFRACTION	99.72
4.28-5.39	0.1684	137	0.1208	5132	X-RAY DIFFRACTION	99.72
5.39-29.69	0.164	137	0.1591	5143	X-RAY DIFFRACTION	99.53