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Literature summary for 5.4.3.5 extracted from

  • Menon, B.R.; Fisher, K.; Rigby, S.E.; Scrutton, N.S.; Leys, D.
    A conformational sampling model for radical catalysis in pyridoxal phosphate- and cobalamin-dependent enzymes (2014), J. Biol. Chem., 289, 34161-34174 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 5.4.3.5

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of wild-type and beta-subunit mutant enzymes Acetoanaerobium sticklandii

Protein Variants

Protein Variants Comment Organism
C700S site-directed mutagenesis, the beta-subunit mutant shows similar kinetics and activity as the wild-type enzyme Acetoanaerobium sticklandii
D627A site-directed mutagenesis, the beta-subunit mutant shows similar kinetics and slightly reduced activity compared to the wild-type enzyme Acetoanaerobium sticklandii
E338A site-directed mutagenesis, the beta-subunit mutant shows altered kinetics and highly reduced activity compared to the wild-type enzyme Acetoanaerobium sticklandii
G128D site-directed mutagenesis, inactive beta-subunit mutant Acetoanaerobium sticklandii
G339W site-directed mutagenesis, the beta-subunit mutant shows altered kinetics and highly reduced activity compared to the wild-type enzyme Acetoanaerobium sticklandii
I424E site-directed mutagenesis, the beta-subunit mutant shows altered kinetics and reduced activity compared to the wild-type enzyme Acetoanaerobium sticklandii
additional information OAM variants are designed to perturb the interface between the cobalamin-binding domain and the pyridoxal 5'-phosphate-binding TIM-barrel domain. Steady-state and single turnover kinetic studies of these variants, combined with pulsed electron-electron double resonance measurements of spin-labeled OAM are used to provide direct evidence for a dynamic interface between the cobalamin and pyridoxal 5'-phosphate-binding domains, overview Acetoanaerobium sticklandii
P343W site-directed mutagenesis, the beta-subunit mutant shows altered kinetics and highly reduced activity compared to the wild-type enzyme Acetoanaerobium sticklandii

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information steady-state and pre-steady-state kinetics Acetoanaerobium sticklandii
0.176
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant E338A Acetoanaerobium sticklandii
0.185
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant I424E Acetoanaerobium sticklandii
0.186
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant C700S Acetoanaerobium sticklandii
0.189
-
 D-ornithine pH 8.5, 25°C, recombinant wild-type enzyme Acetoanaerobium sticklandii
0.19
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant G339W Acetoanaerobium sticklandii
0.193
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant D627A Acetoanaerobium sticklandii
0.193
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant P343W Acetoanaerobium sticklandii

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-ornithine Acetoanaerobium sticklandii
-
 (2R,4S)-2,4-diaminopentanoate
-
 r

Organism

Organism UniProt Comment Textmining
Acetoanaerobium sticklandii E3PY95 AND E3PY96 subunits beta and alpha
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and beta-subunit mutant enzymes Acetoanaerobium sticklandii

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-ornithine
-
 Acetoanaerobium sticklandii (2R,4S)-2,4-diaminopentanoate
-
 r
additional information cobalamin-dependent enzymes enhance the rate of C-Co bond cleavage by up to 1012-fold to generate cob(II)alamin and a transient adenosyl radical. In the case of the pyridoxal 5'-phosphate and cobalamin-dependent enzymes lysine 5,6-aminomutase (EC 5.4.3.3) and ornithine 4,5 aminomutase, it has been proposed that a large scale domain reorientation of the cobalamin-binding domain is linked to radical catalysis Coupled enzyme assay with (2R,4S)-2,4-diaminopentanoate dehydrogenase (DAPDH) from Clostridium difficile Acetoanaerobium sticklandii ?
-
 ?

Synonyms

Synonyms Comment Organism
OAM
-
 Acetoanaerobium sticklandii
ornithine 4,5-aminomutase
-
 Acetoanaerobium sticklandii

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Acetoanaerobium sticklandii

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.14
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant P343W Acetoanaerobium sticklandii
0.2
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant G339W Acetoanaerobium sticklandii
0.24
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant E338A Acetoanaerobium sticklandii
0.76
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant I424E Acetoanaerobium sticklandii
2.88
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant C700S Acetoanaerobium sticklandii
2.89
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant D627A Acetoanaerobium sticklandii
2.97
-
 D-ornithine pH 8.5, 25°C, recombinant wild-type enzyme Acetoanaerobium sticklandii

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8.5
-
 assay at Acetoanaerobium sticklandii

Cofactor

Cofactor Comment Organism Structure
5'-deoxyadenosylcobalamin
-
 Acetoanaerobium sticklandii
additional information ornithine 4,5-aminomutase is a 5'-deoxyadenosylcobalamin and pyridoxal 5'-phosphate co-dependent radical enzyme. Identification of a dynamic interface between the cobalamin and pyridoxal 5'-phosphate-binding domains, overview. Following ligand binding-induced cleavage of the Lys629-pyridoxal 5'-phosphate covalent bond, dynamic motion of the cobalamin-binding domain leads to conformational sampling of the available space. This supports radical catalysis through transient formation of a catalytically competent active state. Crucially, it appears that the formation of the state containing both a substrate/product radical and Co(II) does not restrict cobalamin domain motion Acetoanaerobium sticklandii
pyridoxal 5'-phosphate
-
 Acetoanaerobium sticklandii

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.72
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant P343W Acetoanaerobium sticklandii
1.05
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant G339W Acetoanaerobium sticklandii
1.36
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant E338A Acetoanaerobium sticklandii
4.11
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant I424E Acetoanaerobium sticklandii
14.97
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant D627A Acetoanaerobium sticklandii
15.48
-
 D-ornithine pH 8.5, 25°C, recombinant beta-subunit mutant C700S Acetoanaerobium sticklandii
15.71
-
 D-ornithine pH 8.5, 25°C, recombinant wild-type enzyme Acetoanaerobium sticklandii