BRENDA - Enzyme Database

Role of arg100 in the active site of adenosylcobalamin-dependent glutamate mutase

Xia, L.; Ballou, D.P.; Marsh, E.N.; Biochemistry 43, 3238-3245 (2004)

Data extracted from this reference:

Engineering
EC Number
Amino acid exchange
Commentary
Organism
5.4.99.1
R100K
cob(II)alamin accumulates to a concentration similar to that of the wild-type enzyme, homolysis of the coenzyme is slower by an order of magnitude, compared to wild-type enzyme, glutamate binding is significantly weakened. Mutant does not exhibit the very large deuterium isotope effects that are observed for homolysis of the coenzyme when the wild-type enzyme is reacted with deuterated substrates. Km-value for glutamate is reduced 121fold compared to wild-type enzyme, KM-value for glutamate is increased 17fold compared to wild-type enzyme
Clostridium cochlearium
5.4.99.1
R100M
no cob(II)alamin detected in UV-visible spectrum. Km-value for glutamate is reduced 276fold compared to wild-type enzyme, KM-value for glutamate is increased 13fold compared to wild-type enzyme
Clostridium cochlearium
5.4.99.1
R100Y
no cob(II)alamin detected in UV-visible spectrum. Km-value for glutamate is reduced 322fold compared to wild-type enzyme, KM-value for glutamate is increased 17fold compared to wild-type enzyme
Clostridium cochlearium
KM Value [mM]
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.58
-
L-glutamate
25°C, wild-type enzyme
Clostridium cochlearium
5.4.99.1
7.6
-
L-glutamate
25°C, mutant enzyme R100Y
Clostridium cochlearium
5.4.99.1
10
-
L-glutamate
25°C, mutant enzyme R100K
Clostridium cochlearium
5.4.99.1
13
-
L-glutamate
25°C, mutant enzyme R100M
Clostridium cochlearium
Organism
EC Number
Organism
Primary Accession No. (UniProt)
Commentary
Textmining
5.4.99.1
Clostridium cochlearium
-
-
-
Substrates and Products (Substrate)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
5.4.99.1
L-glutamate
-
650247
Clostridium cochlearium
L-threo-3-methylaspartate
-
650247
Clostridium cochlearium
?
Turnover Number [1/s]
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.0181
-
L-glutamate
25°C, mutant enzyme R100Y
Clostridium cochlearium
5.4.99.1
0.0211
-
L-glutamate
25°C, mutant enzyme R100M
Clostridium cochlearium
5.4.99.1
0.048
-
L-glutamate
25°C, mutant enzyme R100K
Clostridium cochlearium
5.4.99.1
5.8
-
L-glutamate
25°C, wild-type enzyme
Clostridium cochlearium
Cofactor
EC Number
Cofactor
Commentary
Organism
Structure
5.4.99.1
adenosylcobalamin
Km-value for wild-type enzyme is 0.005 mM. No cob(II)alamin detected in UV-visible spectrum of mutant enzymes R100M and R100Y. In mutant enzyme R100K cob(II)alamin accumulates to a concentration similar to that of the wild-type enzyme, homolysis of the coenzyme is slower by an order of magnitude, compared to wild-type enzyme. Mutant does not exhibit the very large deuterium isotope effects that are observed for homolysis of the coenzyme when the wild-type enzyme is reacted with deuterated substrates
Clostridium cochlearium
Cofactor (protein specific)
EC Number
Cofactor
Commentary
Organism
Structure
5.4.99.1
adenosylcobalamin
Km-value for wild-type enzyme is 0.005 mM. No cob(II)alamin detected in UV-visible spectrum of mutant enzymes R100M and R100Y. In mutant enzyme R100K cob(II)alamin accumulates to a concentration similar to that of the wild-type enzyme, homolysis of the coenzyme is slower by an order of magnitude, compared to wild-type enzyme. Mutant does not exhibit the very large deuterium isotope effects that are observed for homolysis of the coenzyme when the wild-type enzyme is reacted with deuterated substrates
Clostridium cochlearium
Engineering (protein specific)
EC Number
Amino acid exchange
Commentary
Organism
5.4.99.1
R100K
cob(II)alamin accumulates to a concentration similar to that of the wild-type enzyme, homolysis of the coenzyme is slower by an order of magnitude, compared to wild-type enzyme, glutamate binding is significantly weakened. Mutant does not exhibit the very large deuterium isotope effects that are observed for homolysis of the coenzyme when the wild-type enzyme is reacted with deuterated substrates. Km-value for glutamate is reduced 121fold compared to wild-type enzyme, KM-value for glutamate is increased 17fold compared to wild-type enzyme
Clostridium cochlearium
5.4.99.1
R100M
no cob(II)alamin detected in UV-visible spectrum. Km-value for glutamate is reduced 276fold compared to wild-type enzyme, KM-value for glutamate is increased 13fold compared to wild-type enzyme
Clostridium cochlearium
5.4.99.1
R100Y
no cob(II)alamin detected in UV-visible spectrum. Km-value for glutamate is reduced 322fold compared to wild-type enzyme, KM-value for glutamate is increased 17fold compared to wild-type enzyme
Clostridium cochlearium
KM Value [mM] (protein specific)
EC Number
KM Value [mM]
KM Value Maximum [mM]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.58
-
L-glutamate
25°C, wild-type enzyme
Clostridium cochlearium
5.4.99.1
7.6
-
L-glutamate
25°C, mutant enzyme R100Y
Clostridium cochlearium
5.4.99.1
10
-
L-glutamate
25°C, mutant enzyme R100K
Clostridium cochlearium
5.4.99.1
13
-
L-glutamate
25°C, mutant enzyme R100M
Clostridium cochlearium
Substrates and Products (Substrate) (protein specific)
EC Number
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
5.4.99.1
L-glutamate
-
650247
Clostridium cochlearium
L-threo-3-methylaspartate
-
650247
Clostridium cochlearium
?
Turnover Number [1/s] (protein specific)
EC Number
Turnover Number Minimum [1/s]
Turnover Number Maximum [1/s]
Substrate
Commentary
Organism
Structure
5.4.99.1
0.0181
-
L-glutamate
25°C, mutant enzyme R100Y
Clostridium cochlearium
5.4.99.1
0.0211
-
L-glutamate
25°C, mutant enzyme R100M
Clostridium cochlearium
5.4.99.1
0.048
-
L-glutamate
25°C, mutant enzyme R100K
Clostridium cochlearium
5.4.99.1
5.8
-
L-glutamate
25°C, wild-type enzyme
Clostridium cochlearium