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Literature summary for 6.3.5.5 extracted from

  • Kim, J.; Raushel, F.M.
    Perforation of the tunnel wall in carbamoyl phosphate synthetase derails the passage of ammonia between sequential active sites (2004), Biochemistry, 43, 5334-5340.
    View publication on PubMed

Protein Variants

Protein Variants Comment Organism
D362A mutation in alpha subunit, 2fold increase in turnover number for NH4+, 5.4fold decrease in KM-value for NH4+, 1.7fold increase in turnover number for Gln, 1.7fold increase in KM-value for Gln Escherichia coli
D362A/betaR265A mutation S362A in alpha-subunit, mutation R265A in beta-subunit,3fold increase in turnover number for NH4+, 2.2fold decrease in KM-value for NH4+, 1.2fold decrease in turnover number for Gln, 71fold increase in KM-value for Gln Escherichia coli
P360A/H361A mutation in beta-subunit, turnover number for NH4+ is nearly identical to wild-type value, 1.6fold increase in turnover number for Gln, 3.2fold increase in KM-value for Gln Escherichia coli
P360A/H361A/R265A mutations P360A and H361A in alpha-subunit, mutation R265A in beta-subunit, mutant enzyme is unable to utilize glutamine for the synthesis of carbamoyl phosphate1.3fold increase in turnover number for NH4+, 11.8fold decrease in KM-value for NH4+ Escherichia coli
Q262A/R265A mutation in beta-subunit 1.9fold increase in turnover number for NH4+, 5fold decrease in KM-value for NH4+, 1.4fold decrease in turnover number for Gln, 43fold increase in KM-value for Gln Escherichia coli
Q262A/R265A/N266A mutation in beta-subunit, 1.6fold decrease in turnover number for Gln, 13.5fold increase in KM-value for Gln Escherichia coli
R265A mutation in beta-subunit, 1.3fold increase in turnover number for NH4+, 5fold decrease in KM-value for NH4+, 1.3fold decrease in turnover number for Gln, 69fold increase in KM-value for Gln Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.11
-
L-Gln pH 7.6, wild-type enzyme Escherichia coli
0.19
-
L-Gln pH 7.6, mutant alphaD362A Escherichia coli
0.36
-
L-Gln pH 7.6, mutant alphaP360A/alphaH361A Escherichia coli
4.7
-
L-Gln pH 7.6, mutant betaQ262A/betaR265A Escherichia coli
7.6
-
L-Gln pH 7.6, mutant betaR265A Escherichia coli
7.8
-
L-Gln pH 7.6, mutant alphaD362A/betaR265A Escherichia coli
12
-
NH4+ pH 7.6, mutant alphaP360A/alphaH361A/betaR265A Escherichia coli
14.9
-
L-Gln pH 7.6, mutant betaQ262A/betaR265A/betaN266A Escherichia coli
24
-
NH4+ pH 7.6, mutant alphaD362A Escherichia coli
26
-
NH4+ pH 7.6, mutant betaQ262A/betaR265A Escherichia coli
26
-
NH4+ pH 7.6, mutant betaR265A Escherichia coli
59
-
NH4+ pH 7.6, mutant alphaD362A/betaR265A Escherichia coli
130
-
NH4+ pH 7.6, wild-type enzyme Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2 ATP + L-Gln + HCO3- carbamoyl phosphate synthetase contains an internal molecular tunnel, which has been proposed to facilitate the translocation of reaction intermediates from one active site to another. Ammonia, the product from the hydrolysis of glutamine in the small subunit, is apparently transported to the next active site in the large subunit of CPS over a distance of about 45 Å. The ammonia tunnel that connects these two active sites provides a direct path for the guided diffusion of ammonia and protection from protonation Escherichia coli 2 ADP + phosphate + L-Glu + carbamoyl phosphate
-
?
2 ATP + NH4+ + HCO3- the enzyme is able to utilize external NH4+ as an alternative nitrogen source when glutamine is absent Escherichia coli 2 ADP + phosphate + carbamoyl phosphate
-
?

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.005
-
L-Gln pH 7.6, mutant alphaP360A/alphaH361A/betaR265A Escherichia coli
0.88
-
L-Gln pH 7.6, mutant betaQ262A/betaR265A/betaN266A Escherichia coli
1
-
L-Gln pH 7.6, mutant betaQ262A/betaR265A Escherichia coli
1.1
-
L-Gln pH 7.6, mutant betaR265A Escherichia coli
1.2
-
L-Gln pH 7.6, mutant alphaD362A/betaR265A Escherichia coli
1.4
-
L-Gln pH 7.6, wild-type enzyme Escherichia coli
1.9
-
NH4+ pH 7.6, wild-type enzyme Escherichia coli
2
-
NH4+ pH 7.6, mutant alphaP360A/alphaH361A Escherichia coli
2.3
-
L-Gln pH 7.6, mutant alphaP360A/alphaH361A Escherichia coli
2.4
-
L-Gln pH 7.6, mutant alphaD362A Escherichia coli
2.5
-
NH4+ pH 7.6, mutant alphaP360A/alphaH361A/betaR265A Escherichia coli
2.5
-
NH4+ pH 7.6, mutant betaR265A Escherichia coli
3.6
-
NH4+ pH 7.6, mutant betaQ262A/betaR265A Escherichia coli
3.8
-
NH4+ pH 7.6, mutant alphaD362A Escherichia coli
5.8
-
NH4+ pH 7.6, mutant alphaD362A/betaR265A Escherichia coli