PDB-7sa8: Crystal Structure of the periplasmic lyase AlgL K66A Mutant

ID or keywords:

Entry

Database: PDB / ID: 7sa8

Title

Crystal Structure of the periplasmic lyase AlgL K66A Mutant

Components

Alginate lyase

Keywords

LYASE / Alginate lyase

Function / homology

Function and homology information

alginic acid catabolic process / poly(beta-D-mannuronate) lyase activity / periplasmic space
Similarity search - Function

Biological species

Pseudomonas aeruginosa (bacteria)

Method

X-RAY DIFFRACTION /

MOLECULAR REPLACEMENT / Resolution: 2.5 Å

Authors

Gheorghita, A.A. / Pfoh, R. / Wong, S.S.Y. / Howell, P.L.

Funding support

Canada, 2items

Citation

Journal: J.Biol.Chem. / Year: 2022
Title: The Pseudomonas aeruginosa homeostasis enzyme AlgL clears the periplasmic space of accumulated alginate during polymer biosynthesis.
Authors: Gheorghita, A.A. / Wolfram, F. / Whitfield, G.B. / Jacobs, H.M. / Pfoh, R. / Wong, S.S.Y. / Guitor, A.K. / Goodyear, M.C. / Berezuk, A.M. / Khursigara, C.M. / Parsek, M.R. / Howell, P.L.

History

Deposition	Sep 22, 2021	Deposition site: RCSB / Processing site: RCSB
Revision 1.0	Aug 3, 2022	Provider: repository / Type: Initial release
Revision 1.1	Oct 18, 2023	Group: Data collection / Refinement description Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.2	Nov 13, 2024	Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature

Structure viewer	Molecule: MolmilJmol/JSmol

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Download

PDBx/mmCIF format	7sa8.cif.gz	128.6 KB	Display	PDBx/mmCIF format
PDB format	pdb7sa8.ent.gz	95.4 KB	Display	PDB format
PDBx/mmJSON format	7sa8.json.gz		Tree view	PDBx/mmJSON format
Others	Other downloads

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Validation report

Summary document	7sa8_validation.pdf.gz	429.2 KB	Display	wwPDB validaton report
Full document	7sa8_full_validation.pdf.gz	433.5 KB	Display
Data in XML	7sa8_validation.xml.gz	13 KB	Display
Data in CIF	7sa8_validation.cif.gz	17.3 KB	Display
Arichive directory	https://data.pdbj.org/pub/pdb/validation_reports/sa/7sa8 ftp://data.pdbj.org/pub/pdb/validation_reports/sa/7sa8	HTTPS FTP

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Related structure data

Related structure data	4ozvSC 4ozwC 1hv6 1qaz 4e1y 4f10 4f13 S: Starting model for refinement C: citing same article (ref.)
Similar structure data	Similarity search - Function & homologyF&H Search Similarity search - FunctionF&H Search Similarity search - HomologyF&H Search

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Links

PDB pages	PDBj / wwPDB / NCBI

Deposited unit

A: Alginate lyase

39.8 kDa, 1 polymers
Search similar-shape structures of this assembly by Omokage search (details)

1

Idetical with deposited unit
defined by author
Evidence: gel filtration

Unit cell

Length a, b, c (Å)	58.736, 67.496, 76.216
Angle α, β, γ (deg.)	90.000, 90.000, 90.000
Int Tables number	18
Space group name H-M	P2₁2₁2

#1: Protein	Alginate lyase Mass: 39820.723 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pseudomonas aeruginosa (bacteria) / Gene: algL / Production host: Escherichia coli (E. coli) / References: UniProt: A0A2U8U7V5
#2: Water	ChemComp-HOH / water Mass: 18.015 Da / Num. of mol.: 38 / Source method: isolated from a natural source / Formula: H₂O
Has protein modification	Y

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Experiment

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

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Sample preparation

Crystal	Density Matthews: 1.9 Å³/Da / Density % sol: 35.16 %
Crystal grow	Temperature: 293 K / Method: vapor diffusion, hanging drop Details: 0.275 M K2SO4, 19% (w/v) PEG 3350, and 0.1 M HEPES pH 6.9. Crystals were cryoprotected by soaking for 10 min with 2 mM mannuronate tetrasaccharide and 20% (v/v) PEG 4000.

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Data collection

Diffraction	Mean temperature: 100 K / Serial crystal experiment: N
Diffraction source	Source: SEALED TUBE / Type: BRUKER D8 QUEST / Wavelength: 1.5406 Å
Detector	Type: BRUKER PHOTON 100 / Detector: CMOS / Date: May 13, 2015
Radiation	Monochromator: M / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelength	Wavelength: 1.5406 Å / Relative weight: 1
Reflection	Resolution: 2.5→20.305 Å / Num. obs: 10903 / % possible obs: 99.3 % / Redundancy: 10.3 % / R_merge(I) obs: 0.098 / Net I/σ(I): 16
Reflection shell	Resolution: 2.5→2.6 Å / R_merge(I) obs: 0.435 / Num. unique obs: 1196 / % possible all: 100

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Processing

Software

Name	Version	Classification
REFMAC	5.8.0267	refinement
PROTEUM PLUS		data reduction
PROTEUM PLUS		data scaling
PHASER		phasing

Refinement

Method to determine structure:

MOLECULAR REPLACEMENT
Starting model: 4OZV

4ozv

Resolution: 2.5→20.305 Å / Cor.coef. F_o:F_c: 0.912 / Cor.coef. F_o:F_c free: 0.878 / SU B: 29.256 / SU ML: 0.295 / Cross valid method: THROUGHOUT / ESU R: 1.643 / ESU R Free: 0.337
Details: Hydrogens have been added in their riding positions

	R_factor	Num. reflection	% reflection
R_free	0.2689	571	5.237 %
R_work	0.2461	10332	-
all	0.247	-	-
obs	-	10332	99.299 %

Solvent computation

Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK BULK SOLVENT

Displacement parameters

B_iso mean: 39.375 Å²

	B_aniso -1	B_aniso -2	B_aniso -3
1-	-0.699 Å²	0 Å²	-0 Å²
2-	-	1.296 Å²	-0 Å²
3-	-	-	-0.597 Å²

Refinement step

Cycle: LAST / Resolution: 2.5→20.305 Å

	Protein	Nucleic acid	Ligand	Solvent	Total
Num. atoms	2432	0	0	38	2470

Refine LS restraints

Refine-ID	Type	Dev ideal	Dev ideal target	Number
X-RAY DIFFRACTION	r_bond_refined_d	0.007	0.013	2518
X-RAY DIFFRACTION	r_bond_other_d	0.001	0.017	2289
X-RAY DIFFRACTION	r_angle_refined_deg	1.503	1.644	3424
X-RAY DIFFRACTION	r_angle_other_deg	1.273	1.58	5249
X-RAY DIFFRACTION	r_dihedral_angle_1_deg	6.151	5	316
X-RAY DIFFRACTION	r_dihedral_angle_2_deg	30.293	21.799	139
X-RAY DIFFRACTION	r_dihedral_angle_3_deg	15.638	15	385
X-RAY DIFFRACTION	r_dihedral_angle_4_deg	17.024	15	19
X-RAY DIFFRACTION	r_chiral_restr	0.066	0.2	316
X-RAY DIFFRACTION	r_gen_planes_refined	0.006	0.02	2917
X-RAY DIFFRACTION	r_gen_planes_other	0.001	0.02	617
X-RAY DIFFRACTION	r_nbd_refined	0.2	0.2	567
X-RAY DIFFRACTION	r_symmetry_nbd_other	0.188	0.2	2104
X-RAY DIFFRACTION	r_nbtor_refined	0.163	0.2	1243
X-RAY DIFFRACTION	r_symmetry_nbtor_other	0.081	0.2	1096
X-RAY DIFFRACTION	r_xyhbond_nbd_refined	0.16	0.2	73
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_other	0.098	0.2	1
X-RAY DIFFRACTION	r_symmetry_nbd_refined	0.125	0.2	11
X-RAY DIFFRACTION	r_nbd_other	0.202	0.2	35
X-RAY DIFFRACTION	r_symmetry_xyhbond_nbd_refined	0.262	0.2	2
X-RAY DIFFRACTION	r_mcbond_it	1.46	3.18	1258
X-RAY DIFFRACTION	r_mcbond_other	1.451	3.177	1257
X-RAY DIFFRACTION	r_mcangle_it	2.494	4.753	1570
X-RAY DIFFRACTION	r_mcangle_other	2.495	4.756	1571
X-RAY DIFFRACTION	r_scbond_it	1.395	3.344	1259
X-RAY DIFFRACTION	r_scbond_other	1.393	3.341	1258
X-RAY DIFFRACTION	r_scangle_it	2.421	4.931	1851
X-RAY DIFFRACTION	r_scangle_other	2.413	4.93	1851
X-RAY DIFFRACTION	r_lrange_it	6.771	58.763	10818
X-RAY DIFFRACTION	r_lrange_other	6.771	58.736	10805

LS refinement shell

Resolution (Å)	R_factor R_free	Num. reflection R_free	R_factor R_work	Num. reflection R_work	Refine-ID	% reflection obs (%)
2.5-2.564	0.31	35	0.327	750	X-RAY DIFFRACTION	100
2.564-2.634	0.289	45	0.321	731	X-RAY DIFFRACTION	100
2.634-2.71	0.289	37	0.321	714	X-RAY DIFFRACTION	100
2.71-2.793	0.329	35	0.299	694	X-RAY DIFFRACTION	100
2.793-2.883	0.381	33	0.273	657	X-RAY DIFFRACTION	99.5671
2.883-2.984	0.224	34	0.265	661	X-RAY DIFFRACTION	99.8563
2.984-3.095	0.243	36	0.247	641	X-RAY DIFFRACTION	100
3.095-3.22	0.295	33	0.255	588	X-RAY DIFFRACTION	100
3.22-3.361	0.278	31	0.249	579	X-RAY DIFFRACTION	100
3.361-3.523	0.407	33	0.255	559	X-RAY DIFFRACTION	99.3289
3.523-3.711	0.332	35	0.264	522	X-RAY DIFFRACTION	98.4099
3.711-3.932	0.205	25	0.242	501	X-RAY DIFFRACTION	97.7695
3.932-4.199	0.193	28	0.203	484	X-RAY DIFFRACTION	99.2248
4.199-4.528	0.274	36	0.195	433	X-RAY DIFFRACTION	99.5754
4.528-4.949	0.252	26	0.182	403	X-RAY DIFFRACTION	99.7674
4.949-5.514	0.238	29	0.231	372	X-RAY DIFFRACTION	99.5037
5.514-6.332	0.299	12	0.242	357	X-RAY DIFFRACTION	100
6.332-7.67	0.118	14	0.236	294	X-RAY DIFFRACTION	100
7.67-10.509	0.179	10	0.186	247	X-RAY DIFFRACTION	100
10.509-20.305	0.302	4	0.304	143	X-RAY DIFFRACTION	83.5227

Refinement TLS params.

Method: refined / Origin x: 9.039 Å / Origin y: 82.556 Å / Origin z: 19.965 Å

	11	12	13	21	22	23	31	32	33
T	0.0054 Å²	0.0009 Å²	0.0056 Å²	-	0.0294 Å²	-0.0304 Å²	-	-	0.1793 Å²
L	1.0451 °²	-0.2083 °²	0.1033 °²	-	0.187 °²	-0.2679 °²	-	-	1.4904 °²
S	0.033 Å °	0.1573 Å °	-0.2099 Å °	0.0079 Å °	-0.0499 Å °	0.0179 Å °	0.0157 Å °	-0.013 Å °	0.0169 Å °

Refinement TLS group

Selection: ALL

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