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- PDB-1nj4: Crystal structure of a deacylation-defective mutant of penicillin... -

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Basic information

Entry
Database: PDB / ID: 1nj4
TitleCrystal structure of a deacylation-defective mutant of penicillin-binding protein 5 at 1.9 A resolution
ComponentsPenicillin-binding protein 5
KeywordsHYDROLASE / peptidoglycan synthesis / penicllin-binding protein / DD-carboxypeptidase
Function / homology
Function and homology information


peptidoglycan metabolic process / serine-type D-Ala-D-Ala carboxypeptidase / serine-type D-Ala-D-Ala carboxypeptidase activity / penicillin binding / peptidoglycan biosynthetic process / carboxypeptidase activity / cell wall organization / beta-lactamase activity / beta-lactamase / outer membrane-bounded periplasmic space ...peptidoglycan metabolic process / serine-type D-Ala-D-Ala carboxypeptidase / serine-type D-Ala-D-Ala carboxypeptidase activity / penicillin binding / peptidoglycan biosynthetic process / carboxypeptidase activity / cell wall organization / beta-lactamase activity / beta-lactamase / outer membrane-bounded periplasmic space / regulation of cell shape / protein homodimerization activity / proteolysis / plasma membrane
Similarity search - Function
Peptidoglycan synthesis regulatory factor (PBP3), Domain 2 / D-Ala-D-Ala carboxypeptidase, C-terminal domain / D-Ala-D-Ala carboxypeptidase, C-terminal domain superfamily / Peptidase S11, D-Ala-D-Ala carboxypeptidase A, C-terminal / Penicillin-binding protein 5, C-terminal domain / Penicillin-binding protein 5, C-terminal domain / Penicillin-binding protein, C-terminal domain superfamily / Peptidase S11, D-alanyl-D-alanine carboxypeptidase A, N-terminal / Peptidase S11, D-alanyl-D-alanine carboxypeptidase A / D-alanyl-D-alanine carboxypeptidase ...Peptidoglycan synthesis regulatory factor (PBP3), Domain 2 / D-Ala-D-Ala carboxypeptidase, C-terminal domain / D-Ala-D-Ala carboxypeptidase, C-terminal domain superfamily / Peptidase S11, D-Ala-D-Ala carboxypeptidase A, C-terminal / Penicillin-binding protein 5, C-terminal domain / Penicillin-binding protein 5, C-terminal domain / Penicillin-binding protein, C-terminal domain superfamily / Peptidase S11, D-alanyl-D-alanine carboxypeptidase A, N-terminal / Peptidase S11, D-alanyl-D-alanine carboxypeptidase A / D-alanyl-D-alanine carboxypeptidase / Beta-lactamase / DD-peptidase/beta-lactamase superfamily / Beta-lactamase/transpeptidase-like / Sandwich / 3-Layer(aba) Sandwich / Mainly Beta / Alpha Beta
Similarity search - Domain/homology
D-alanyl-D-alanine carboxypeptidase DacA / D-alanyl-D-alanine carboxypeptidase DacA
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodX-RAY DIFFRACTION / Refinement from a lower resolution structure / Resolution: 1.9 Å
AuthorsNicola, G. / Nicholas, R.A. / Davies, C.
Citation
Journal: J.Biol.Chem. / Year: 2003
Title: Crystal structure of wild-type penicillin-binding protein 5 from Escherichia coli: implications for deacylation of the acyl-enzyme complex.
Authors: Nicholas, R.A. / Krings, S. / Tomberg, J. / Nicola, G. / Davies, C.
#1: Journal: J.Biol.Chem. / Year: 2001
Title: Crystal structure of a deacylation-defective mutant of penicllin-binding protein at 2.3 A resolution
Authors: Davies, C. / White, S.W. / Nicholas, R.A.
#2: Journal: Rev.Infect.Dis. / Year: 1988
Title: Relations between beta-lactamases and penicllin-binding proteins: beta-lactamase activity of penicillin-binding protein 5 from Escherichia coli
Authors: Nicholas, R.A. / Strominger, J.L.
#3: Journal: FEBS Lett. / Year: 1984
Title: An amino acid substitution that blocks the deacylation step in the enzyme mechanism of penicillin-binding protein 5 of Escherichia coli
Authors: Broome-Smith, J. / Spratt, B.G.
History
DepositionDec 30, 2002Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 7, 2003Provider: repository / Type: Initial release
Revision 1.1Apr 29, 2008Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Version format compliance
Revision 1.3Oct 27, 2021Group: Database references / Category: database_2 / struct_ref_seq_dif
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ref_seq_dif.details
Revision 1.4Aug 16, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Remark 999SEQUENCE This is a soluble construct of a mutant PBP 5, termed sPBP 5'. To produce sPBP5', codons ...SEQUENCE This is a soluble construct of a mutant PBP 5, termed sPBP 5'. To produce sPBP5', codons corresponding to the last 17 amino acid residues were removed but an additional six amino acids (GDPVID) were added due to read through to the stop codon. None of these non-native residues are visible in the electron density map. The first 29 amino acids of the protein encoded by the open reading frame represent the signal sequence, which is removed during maturation and transport to the periplasmic space. These residues are not present in this construct.

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Penicillin-binding protein 5


Theoretical massNumber of molelcules
Total (without water)39,8991
Polymers39,8991
Non-polymers00
Water3,945219
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)50.20, 50.20, 135.76
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number145
Space group name H-MP32

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Components

#1: Protein Penicillin-binding protein 5 / D-alanyl-D-alanine carboxypeptidase fraction A / DD-peptidase / DD-carboxypeptidase / PBP-5


Mass: 39899.152 Da / Num. of mol.: 1 / Mutation: G105D
Source method: isolated from a genetically manipulated source
Details: This mutant form of PBP 5 is termed sPBP 5' / Source: (gene. exp.) Escherichia coli (E. coli) / Gene: DACA / Plasmid: PBR322 / Production host: Escherichia coli (E. coli) / Strain (production host): MC1061
References: UniProt: P04287, UniProt: P0AEB2*PLUS, serine-type D-Ala-D-Ala carboxypeptidase
#2: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 219 / Source method: isolated from a natural source / Formula: H2O

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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: 2.48 Å3/Da / Density % sol: 50.31 %
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7
Details: 20% PEG 4000, 100 mM Tris pH 7.0, VAPOR DIFFUSION, HANGING DROP, temperature 293K
Crystal grow
*PLUS
pH: 7.5
Components of the solutions
*PLUS
IDConc.Common nameCrystal-IDSol-IDDetailsChemical formula
15.5 mg/mlprotein1drop
220 mMTris1droppH7.5
330 mM1dropNaCl
43 mMbeta-mercaptoethanol1drop
520 %PEG40001reservoir
650 mMTris1reservoirpH7.0

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: ROTATING ANODE / Type: RIGAKU RU300 / Wavelength: 1.5418 Å
DetectorType: RIGAKU RAXIS IV / Detector: IMAGE PLATE / Date: Oct 8, 2002 / Details: mirrors
RadiationMonochromator: Osmic mirrors / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.5418 Å / Relative weight: 1
ReflectionResolution: 1.9→36.6 Å / Num. obs: 29772 / % possible obs: 98.7 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 6 % / Biso Wilson estimate: 25.9 Å2 / Rmerge(I) obs: 0.073 / Net I/σ(I): 7.1
Reflection shellResolution: 1.9→1.97 Å / Redundancy: 3.4 % / Rmerge(I) obs: 0.354 / Mean I/σ(I) obs: 1.8 / Num. unique all: 3024 / % possible all: 88.4
Reflection
*PLUS
Num. measured all: 178730
Reflection shell
*PLUS
% possible obs: 88.4 % / Rmerge(I) obs: 0.358

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Processing

Software
NameVersionClassification
REFMAC5refinement
CrystalClear(MSC/RIGAKU)data reduction
d*TREKdata scaling
RefinementMethod to determine structure: Refinement from a lower resolution structure
Starting model: PDB ENTRY 1HD8

1hd8


Resolution: 1.9→15 Å / Cor.coef. Fo:Fc: 0.949 / Cor.coef. Fo:Fc free: 0.927 / SU B: 4.413 / SU ML: 0.127 / Isotropic thermal model: isotropic / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.166 / ESU R Free: 0.151 / Stereochemistry target values: MAXIMUM LIKELIHOOD
RfactorNum. reflection% reflectionSelection details
Rfree0.24454 1507 5.1 %RANDOM
Rwork0.20727 ---
all0.2091 29705 --
obs0.20913 29705 98.7 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: BABINET MODEL WITH MASK
Displacement parametersBiso mean: 29.263 Å2
Baniso -1Baniso -2Baniso -3
1--0.34 Å2-0.17 Å20 Å2
2---0.34 Å20 Å2
3---0.51 Å2
Refinement stepCycle: LAST / Resolution: 1.9→15 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2649 0 0 219 2868
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0070.0222693
X-RAY DIFFRACTIONr_bond_other_d
X-RAY DIFFRACTIONr_angle_refined_deg1.2121.963633
X-RAY DIFFRACTIONr_angle_other_deg
X-RAY DIFFRACTIONr_dihedral_angle_1_deg3.3323341
X-RAY DIFFRACTIONr_dihedral_angle_3_deg18.48315500
X-RAY DIFFRACTIONr_chiral_restr0.0850.2406
X-RAY DIFFRACTIONr_gen_planes_refined0.0040.022019
X-RAY DIFFRACTIONr_gen_planes_other
X-RAY DIFFRACTIONr_nbd_refined0.2240.31246
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined0.1690.5304
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined0.1770.336
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined0.1070.55
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_mcbond_it0.621.51694
X-RAY DIFFRACTIONr_mcangle_it1.18722716
X-RAY DIFFRACTIONr_scbond_it1.6333999
X-RAY DIFFRACTIONr_scangle_it2.7574.5917
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.9→1.949 Å / Total num. of bins used: 20 /
RfactorNum. reflection
Rfree0.371 90
Rwork0.359 1817
Refinement
*PLUS
Lowest resolution: 15 Å / Rfactor obs: 0.209 / Rfactor Rfree: 0.244 / Rfactor Rwork: 0.207
Solvent computation
*PLUS
Displacement parameters
*PLUS
Refine LS restraints
*PLUS
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONr_bond_d0.007
X-RAY DIFFRACTIONr_angle_d1.21

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