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Literature summary extracted from

  • Schnicker, N.; Dey, M.
    Bacillus anthracis prolyl 4-hydroxylase modifies collagen-like substrates in asymmetric patterns (2016), J. Biol. Chem., 291, 13360-13374 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.11.2 gene bap4h, recombinant expression in Escherichia coli strain BL21 star DE3 pLysS cells Bacillus anthracis

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.14.11.2 purified recombinant apoenzyme in complex with inhibitor malonate and Co2+, hanging drop vapor diffusion, mixing of 0.001 ml of a solution of 9 mg/ml protein, 1 mM CoCl2, and 1 mM L-Pro, with 0.001 ml of reservoir solution containing 0.1 M malonate/imidazole/boric acid, pH 6.5, and 18% PEG 1500, 2 days, or purified recombinant apoenzyme BaP4H with (P-P-G)3 peptide fused to the C terminus and Co2+, sitting drop vapor diffusion, mixing of 0.001 ml of 12 mg/ml protein solution containing 1 mM CoCl2, and 1 mM 2-oxoglutarate, with 0.001 ml of reservoir solution containing 0.15 M KBr and 30% PEG 2000 MME, 20°C, method optimization, X-ray diffraction structure determination and analysis, molecular replacement for apo-BaP4H using structure PDB ID 3ITQ as a starting model. Models for Co(II)-BaP4H-MLI and Co(II)-BaP4H-PPG are obtained by molecular replacement with the apo-BaP4H structure Bacillus anthracis

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.14.11.2 Co2+ active site binding structure analysis, in both Co(II)-bound structures, in addition to the catalytic triad residues and 2-oxoglutarate or, malonate that coordinate cobalt, the remaining coordinate sites are occupied by water molecules thus forming a six-coordinate cobalt complex Bacillus anthracis
1.14.11.2 malonate active site binding structure analysis Bacillus anthracis

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.14.11.2 Fe2+ required for catalysis, 1:1 ratio of Fe(II) to BaP4H monomer Bacillus anthracis

Organism

EC Number Organism UniProt Comment Textmining
1.14.11.2 Bacillus anthracis Q81LZ8
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.11.2 recombinant enzyme from Escherichia coli strain BL21 star DE3 pLysS cells Bacillus anthracis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.11.2 human procollagen-like (P-P-G)10 peptide L-proline + 2-oxoglutarate + O2 4 different peptide sequences, anaerobic conditions Bacillus anthracis human procollagen-like (P-P-G)10 peptide trans-4-hydroxy-L-proline + succinate + CO2
-
 ?
1.14.11.2 human procollagen-like (P-P-G)5 peptide L-proline + 2-oxoglutarate + O2 5 different peptide sequences, anaerobic conditions Bacillus anthracis human procollagen-like (P-P-G)5 peptide trans-4-hydroxy-L-proline + succinate + CO2
-
 ?
1.14.11.2 additional information enzyme BaP4H recognizes and acts on peptidyl substrates but not free L-proline, using elements characteristic of an Fe(II)/2-oxoglutarate-dependent dioxygenases, substrate specifiicty analysis by mass spectrometry, fluorescence binding, x-ray crystallography, and docking experiments, overview. Enzyme BaP4H can hydroxylate unique peptidyl proline sites in collagen-derived peptides with asymmetric hydroxylation patterns. The cofactor-bound crystal structures of BaP4H reveal active site conformational changes that define open and closed forms and mimic 'ready' and 'product-released' states of the enzyme in the catalytic cycle. Free L-proline binds to Fe(II)-BaP4H, but the affinity is an order of magnitude lower than for the peptides. No activity with free L-proline Bacillus anthracis ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
1.14.11.2 BaP4H
-
 Bacillus anthracis
1.14.11.2 P4H
-
 Bacillus anthracis
1.14.11.2 prolyl 4-hydroxylase
-
 Bacillus anthracis

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.14.11.2 20
-
 assay at Bacillus anthracis

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.14.11.2 7.4
-
 assay at Bacillus anthracis

Cofactor

EC Number Cofactor Comment Organism Structure
1.14.11.2 ascorbate
-
 Bacillus anthracis

General Information

EC Number General Information Comment Organism
1.14.11.2 additional information ligand docking calculations, substrate and inhibitor binding structure, structure-function analysis, coformations of key active site residues Tyr118, Tyr124, and Phe160 in apoenzyme and ligand-bound enzyme, conformational changes involving residues Tyr118, Tyr124, and Phe160 upon metal and 2-oxoglutarate or malonate binding, detailed overview Bacillus anthracis