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Literature summary for 2.4.2.21 extracted from

  • Claas, K.R.; Parrish, J.R.; Maggio-Hall, L.A.; Escalante-Semerena, J.C.
    Functional analysis of the nicotinate mononucleotide:5,6-dimethylbenzimidazole phosphoribosyltransferase (CobT) enzyme, involved in the late steps of coenzyme B12 biosynthesis in Salmonella enterica (2010), J. Bacteriol., 192, 145-154.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 2.4.2.21

Cloned(Commentary)

Cloned (Comment) Organism
wild-type and mutants cloned into vector pSU18 or vector pT7-5. Plasmids carrying wild-type or mutant cobT alleles introduced into strain JE2607 (cobT cobB recA) or strain JE2423 (cobT recA). Plasmids encoding mutant cobT alleles under the control of the T7 promoter of vector pT7-5 transformed into strain JE2017 and overexpressed Salmonella enterica

Protein Variants

Protein Variants Comment Organism
C160A loss of catalytic efficiency by 79%. Renders the enzyme 5fold less efficient than wild-type. Displays no discernible growth defect in no-carbon essential (NCE) medium containing dicyanocobinamide and glycerol or ethanolamine relative to that of a strain that synthesizes wild-type Salmonella enterica
G171D loss of catalytic efficiency by more than 99%, has less affinity for 5,6-dimethylbenzimidazole. Retains activity to support growth, but only when 5,6-dimethylbenzimidazole is present in the medium. Does not support the growth of strain JE2423 on ethanolamine in medium that contains dicyanocobinamide but lacks 5,6-dimethylbenzimidazole Salmonella enterica
G171D/M325A requires cobalamin for growth regardless of the carbon source used Salmonella enterica
G257A loss of catalytic efficiency by 64% Salmonella enterica
G257D loss of catalytic efficiency by more than 99%, has less affinity for 5,6-dimethylbenzimidazole. Does not respond to 5,6-dimethylbenzimidazole and requires cobalamin to grow. Does not support the growth of strain JE2423 on ethanolamine in medium that contains dicyanocobinamide but lacks 5,6-dimethylbenzimidazole Salmonella enterica
G257D/P12G is cobalamin auxotroph on glycerol or ethanolamine. P12G does not alter the effect of the G257D mutation Salmonella enterica
G320D loss of catalytic efficiency by more than 99%, is unstable, retains activity to support growth, but only when 5,6-dimethylbenzimidazole is present in the medium. Does not support the growth of strain JE2423 on ethanolamine in medium that contains dicyanocobinamide but lacks 5,6-dimethylbenzimidazole Salmonella enterica
H289A counteracts the effect of the G320D substitution Salmonella enterica
M325A grows in the absence of exogenous 5,6-dimethylbenzimidazole, albeit at a rate slower than that of a strain making wild-type. Addition of 5,6-dimethylbenzimidazole to the medium allows the strain making M325A to grow as fast as a strain making wild-type Salmonella enterica
P12G does not require exogenous 5,6-dimethylbenzimidazole for growth on glycerol or ethanolamine Salmonella enterica

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.3
-
 beta-nicotinate D-ribonucleotide mutant G257A, at 37°C Salmonella enterica
0.3
-
 beta-nicotinate D-ribonucleotide wild-type, at 37°C Salmonella enterica
0.9
-
 beta-nicotinate D-ribonucleotide mutant G171D, at 37°C Salmonella enterica
1
-
 beta-nicotinate D-ribonucleotide mutant G320D, at 37°C Salmonella enterica
1.2
-
 beta-nicotinate D-ribonucleotide mutant C160A, at 37°C Salmonella enterica
2.2
-
 beta-nicotinate D-ribonucleotide mutant G257D, at 37°C Salmonella enterica

Organism

Organism UniProt Comment Textmining
Salmonella enterica Q05603
-
-

Purification (Commentary)

Purification (Comment) Organism
by gel filtration Salmonella enterica

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.1
-
 mutant G257D, at 37°C Salmonella enterica
0.2
-
 mutant G320D, at 37°C Salmonella enterica
0.5
-
 mutant G171D, at 37°C Salmonella enterica
11
-
 mutant G257A, at 37°C Salmonella enterica
25
-
 mutant C160A, at 37°C Salmonella enterica
30
-
 wild-type, at 37°C Salmonella enterica

Storage Stability

Storage Stability Organism
-80°C, 50 mM Tris-HCl buffer, pH 7.5, 3 M glycerol Salmonella enterica

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
beta-nicotinate D-ribonucleotide + 5,6-dimethylbenzimidazole
-
 Salmonella enterica nicotinate + N1-(5-phospho-alpha-D-ribosyl)-5,6-dimethylbenzimidazole
-
 ?

Synonyms

Synonyms Comment Organism
5,6-dimethylbenzimidazole phosphoribosyltransferase
-
 Salmonella enterica
CobT
-
 Salmonella enterica

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.2
-
 beta-nicotinate D-ribonucleotide mutant G257D, at 37°C Salmonella enterica
0.2
-
 beta-nicotinate D-ribonucleotide mutant G320D, at 37°C Salmonella enterica
0.6
-
 beta-nicotinate D-ribonucleotide mutant G171D, at 37°C Salmonella enterica
14
-
 beta-nicotinate D-ribonucleotide mutant G257A, at 37°C Salmonella enterica
31
-
 beta-nicotinate D-ribonucleotide mutant C160A, at 37°C Salmonella enterica
38
-
 beta-nicotinate D-ribonucleotide wild-type, at 37°C Salmonella enterica

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.1
-
 beta-nicotinate D-ribonucleotide mutant G257D, at 37°C Salmonella enterica
0.2
-
 beta-nicotinate D-ribonucleotide mutant G320D, at 37°C Salmonella enterica
0.7
-
 beta-nicotinate D-ribonucleotide mutant G171D, at 37°C Salmonella enterica
29
-
 beta-nicotinate D-ribonucleotide mutant C160A, at 37°C Salmonella enterica
50
-
 beta-nicotinate D-ribonucleotide mutant G257A, at 37°C Salmonella enterica
138
-
 beta-nicotinate D-ribonucleotide wild-type, at 37°C Salmonella enterica