Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 2.5.1.17 extracted from

  • Mera, P.; Maurice, M.; Rayment, I.; Escalante-Semerena, J.
    Residue Phe112 of the human-type corrinoid adenosyltransferase (PduO) enzyme of lactobacillus reuteri is critical to the formation of the four-coordinate Co(II) corrinoid substrate and to the activity of the enzyme (2009), Biochemistry, 48, 3138-3145.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 2.5.1.17

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli as an N-terminal His-tagged fusion protein Limosilactobacillus reuteri

Protein Variants

Protein Variants Comment Organism
DELTAS183 Km (mM) (Co+ assay): 0.0031 (ATP), 0.00007 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.023 (ATP), 0.02 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0099 (ATP), 0.0163 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.017 (ATP), 0.018 (cob(II)alamin) Limosilactobacillus reuteri
F112A Km (mM) (Co+ assay): 0.0346 (ATP), 0.0019 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0056 (ATP), 0.006 (cob(I)alamin). Mutant F112A shows no activity in the Co2+ assay. The crystal structure of mutant F112A reveal that cob(II)alamin binds to the active site as a five-coordinate species with 5,6-dimethylbenzimidazole serving as a fifth axial ligand. In the Co+ assay [adenosylation of cob(I)alamin], mutant F112A is catalytically competent and displays only a slight decrease in kcat, supposing that in the absence of a four-coordinate cob(II)alamin species, the enzyme is inactive Limosilactobacillus reuteri
F112H Km (mM) (Co+ assay): 0.0026 (ATP), 0.0028 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.00038 (ATP), 0.0042 (cob(I)alamin), Km (mM) (Co2+ assay): 0.01 (ATP), 0.053 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.00059 (ATP), 0.00067 (cob(II)alamin). The crystal structure of an mutantF112H variant show a 5,6-dimethylbenzimidazole-off/His-on interaction between the corrinoid and the enzyme, whose catalytic efficiency is 4 orders of magnitude lower than that of the wild-type protein. The analysis of the kinetic parameters of mutantF112H suggests that the F112H substitution negatively impacts product release Limosilactobacillus reuteri
F112W Km (mM) (Co+ assay): 0.002 (ATP), 0.00077 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0018 (ATP), 0.002 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0073 (ATP), 0.028 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.0028 (ATP), 0.0032 (cob(II)alamin) Limosilactobacillus reuteri
F112Y Km (mM) (Co+ assay): 0.0012 (ATP), 0.00055 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.0016 (ATP), 0.0017 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0094 (ATP), 0.034 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.0023 (ATP), 0.0027 (cob(II)alamin) Limosilactobacillus reuteri
F163A Km (mM) (Co+ assay): 0.0159 (ATP), 0.00019 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.015 (ATP), 0.015 (cob(I)alamin), Km (mM) (Co2+ assay): 0.096 (ATP), 0.134 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.015 (ATP), 0.027 (cob(II)alamin) Limosilactobacillus reuteri
F187A Km (mM) (Co+ assay): 0.0012 (ATP), 0.00011 (cob(I)alamin), kcat (1/sec) (Co+ assay): 0.021 (ATP), 0.018 (cob(I)alamin), Km (mM) (Co2+ assay): 0.0079 (ATP), 0.0158 (cob(II)alamin), kcat (1/sec) (Co+ assay): 0.014 (ATP), 0.015 (cob(II)alamin) Limosilactobacillus reuteri

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.00007
-
 cob(I)alamin mutant DELTAS183, Co+ assay Limosilactobacillus reuteri
0.00011
-
 cob(I)alamin mutant F187A, Co+ assay Limosilactobacillus reuteri
0.00013
-
 cob(I)alamin wild-type, Co+ assay Limosilactobacillus reuteri
0.00019
-
 cob(I)alamin mutant F163A, Co+ assay Limosilactobacillus reuteri
0.00055
-
 cob(I)alamin mutant F112Y, Co+ assay Limosilactobacillus reuteri
0.00077
-
 cob(I)alamin mutant F112W, Co+ assay Limosilactobacillus reuteri
0.0012
-
 ATP mutant F112Y, Co+ assay Limosilactobacillus reuteri
0.0012
-
 ATP mutant F187A, Co+ assay Limosilactobacillus reuteri
0.0019
-
 cob(I)alamin mutant F112A, Co+ assay Limosilactobacillus reuteri
0.002
-
 ATP mutant F112W, Co+ assay Limosilactobacillus reuteri
0.0022
-
 ATP wild-type, Co+ assay Limosilactobacillus reuteri
0.0026
-
 ATP mutant F112H, Co+ assay Limosilactobacillus reuteri
0.0028
-
 cob(I)alamin mutant F112H, Co+ assay Limosilactobacillus reuteri
0.0031
-
 ATP mutant DELTAS183, Co+ assay Limosilactobacillus reuteri
0.005
-
 ATP wild-type, Co2+ assay Limosilactobacillus reuteri
0.0078
-
 cob(II)alamin wild-type, Co2+ assay Limosilactobacillus reuteri
0.0079
-
 ATP mutant F187A, Co2+ assay Limosilactobacillus reuteri
0.0099
-
 ATP mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
0.0104
-
 ATP mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.015
-
 cob(II)alamin mutant F187A, Co2+ assay Limosilactobacillus reuteri
0.0159
-
 ATP mutant F163A, Co+ assay Limosilactobacillus reuteri
0.016
-
 cob(II)alamin mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
0.028
-
 cob(II)alamin mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.034
-
 cob(II)alamin mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.0346
-
 ATP mutant F112A, Co+ assay Limosilactobacillus reuteri
0.053
-
 cob(II)alamin mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.073
-
 ATP mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.094
-
 ATP mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.0961
-
 ATP mutant F163A, Co2+ assay Limosilactobacillus reuteri
0.134
-
 cob(II)alamin mutant F163A, Co2+ assay Limosilactobacillus reuteri

Organism

Organism UniProt Comment Textmining
Limosilactobacillus reuteri
-
-
-

Purification (Commentary)

Purification (Comment) Organism
using Ni-NTA chromatography Limosilactobacillus reuteri

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
cob(I)alamin + ATP in the co+ assay the cobalt ion of cobalamin is chemically reduced in solution to cob(I)alamin by Ti(III)citrate, allowing the cob(I)alamin adenosylation reaction to be measured directly. The Co+ assay is performed under anoxic conditions Limosilactobacillus reuteri adenosylcobalamin + ?
-
 ?
cob(II)alamin + ATP in the Co2+ assay the NADPH-dependent flavodoxin protein reductase/flavodoxin system is used to reduce Co2+ to Co+. In this Co2+ assay, the PduO enzyme must bind cob(II)alamin and facilitate the generation of cob(I)alamin in its active site Limosilactobacillus reuteri adenosylcobalamin + ?
-
 ?

Synonyms

Synonyms Comment Organism
LrPduO
-
 Limosilactobacillus reuteri

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Limosilactobacillus reuteri

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.00038
-
 ATP mutant F112H, Co+ assay Limosilactobacillus reuteri
0.00042
-
 cob(I)alamin mutant F112H, Co+ assay Limosilactobacillus reuteri
0.0005
-
 ATP mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.00067
-
 cob(II)alamin mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.0016
-
 ATP mutant F112Y, Co+ assay Limosilactobacillus reuteri
0.0017
-
 cob(I)alamin mutant F112Y, Co+ assay Limosilactobacillus reuteri
0.0018
-
 ATP mutant F112W, Co+ assay Limosilactobacillus reuteri
0.002
-
 cob(I)alamin mutant F112W, Co+ assay Limosilactobacillus reuteri
0.0023
-
 ATP mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.0027
-
 cob(II)alamin mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.0028
-
 ATP mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.0032
-
 cob(II)alamin mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.0056
-
 ATP mutant F112A, Co+ assay Limosilactobacillus reuteri
0.0066
-
 cob(I)alamin mutant F112A, Co+ assay Limosilactobacillus reuteri
0.014
-
 ATP mutant F187A, Co2+ assay Limosilactobacillus reuteri
0.015
-
 ATP mutant F163A, Co+ assay Limosilactobacillus reuteri
0.015
-
 cob(I)alamin mutant F163A, Co+ assay Limosilactobacillus reuteri
0.015
-
 ATP mutant F163A, Co2+ assay Limosilactobacillus reuteri
0.015
-
 cob(II)alamin mutant F187A, Co2+ assay Limosilactobacillus reuteri
0.017
-
 ATP mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
0.018
-
 cob(II)alamin mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
0.018
-
 cob(I)alamin mutant F187A, Co+ assay Limosilactobacillus reuteri
0.02
-
 cob(I)alamin mutant DELTAS183, Co+ assay Limosilactobacillus reuteri
0.021
-
 ATP mutant F187A, Co+ assay Limosilactobacillus reuteri
0.023
-
 ATP mutant DELTAS183, Co+ assay Limosilactobacillus reuteri
0.024
-
 cob(I)alamin wild-type, Co+ assay Limosilactobacillus reuteri
0.026
-
 ATP wild-type, Co+ assay Limosilactobacillus reuteri
0.027
-
 cob(II)alamin mutant F163A, Co2+ assay Limosilactobacillus reuteri
0.029
-
 ATP wild-type, Co2+ assay Limosilactobacillus reuteri
0.038
-
 cob(II)alamin wild-type, Co2+ assay Limosilactobacillus reuteri

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 assay at Limosilactobacillus reuteri

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.013
-
 cob(II)alamin mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.057
-
 ATP mutant F112H, Co2+ assay Limosilactobacillus reuteri
0.083
-
 cob(II)alamin mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.11
-
 cob(II)alamin mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.14
-
 ATP mutant F112H, Co+ assay Limosilactobacillus reuteri
0.16
-
 ATP mutant F163A, Co2+ assay Limosilactobacillus reuteri
0.17
-
 ATP mutant F112A, Co+ assay Limosilactobacillus reuteri
0.2
-
 cob(II)alamin mutant F163A, Co2+ assay Limosilactobacillus reuteri
0.25
-
 ATP mutant F112Y, Co2+ assay Limosilactobacillus reuteri
0.39
-
 ATP mutant F112W, Co2+ assay Limosilactobacillus reuteri
0.93
-
 cob(II)alamin mutant F187A, Co2+ assay Limosilactobacillus reuteri
0.98
-
 ATP mutant F112W, Co+ assay Limosilactobacillus reuteri
1.1
-
 cob(II)alamin mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
1.4
-
 ATP mutant F112Y, Co+ assay Limosilactobacillus reuteri
1.7
-
 ATP mutant DELTAS183, Co2+ assay Limosilactobacillus reuteri
1.8
-
 ATP mutant F163A, Co+ assay Limosilactobacillus reuteri
1.8
-
 ATP mutant F187A, Co2+ assay Limosilactobacillus reuteri
2.6
-
 cob(I)alamin mutant F112W, Co+ assay Limosilactobacillus reuteri
3.1
-
 cob(I)alamin mutant F112Y, Co+ assay Limosilactobacillus reuteri
3.6
-
 cob(I)alamin mutant F112A, Co+ assay Limosilactobacillus reuteri
4.8
-
 cob(II)alamin wild-type, Co2+ assay Limosilactobacillus reuteri
5.5
-
 ATP wild-type, Co2+ assay Limosilactobacillus reuteri
7.5
-
 ATP mutant DELTAS183, Co+ assay Limosilactobacillus reuteri
12
-
 ATP wild-type, Co+ assay Limosilactobacillus reuteri
15
-
 cob(I)alamin mutant F112H, Co+ assay Limosilactobacillus reuteri
17
-
 ATP mutant F187A, Co+ assay Limosilactobacillus reuteri
79
-
 cob(I)alamin mutant F163A, Co+ assay Limosilactobacillus reuteri
160
-
 cob(I)alamin mutant F187A, Co+ assay Limosilactobacillus reuteri
180
-
 cob(I)alamin wild-type, Co+ assay Limosilactobacillus reuteri
395
-
 cob(I)alamin mutant DELTAS183, Co+ assay Limosilactobacillus reuteri