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Literature summary for 4.6.1.1 extracted from

  • Linder, J.U.
    Structure-function relationships in Escherichia coli adenylate cyclase (2008), Biochem. J., 415, 449-454.
    View publication on PubMed
Search for more literature for 4.6.1.1

Cloned(Commentary)

Cloned (Comment) Organism
class I AC gene, overexpression of wild-type and mutant catalytic domain, residues 2-446, i.e. Cya2-446, in strain BL21(DE3) Escherichia coli

Protein Variants

Protein Variants Comment Organism
D114A site-directed mutagenesis, the almost inactive mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
D116A site-directed mutagenesis, the almost inactive mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
D300A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
E185A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
E242A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
K136A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
K253A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
K260A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
K264A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
K332A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
R19A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
S103A site-directed mutagenesis, the mutant has a 17fold higher Km for ATP compared to the wild-type enzyme, and the mutation causes a marked reduction of discrimination between ATP- and ADP- or AMP-derived inhibitors Escherichia coli
S106A site-directed mutagenesis, the mutation reduces the mutant activity to 25% of the wild-type enzyme activity, kinetic analysis show a 58% reduction of the Vmax and a doubling of the Km compared to the wild-type enzyme Escherichia coli
S113A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
T189A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
W118A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
W200A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Escherichia coli
W249A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
W374A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli
Y394A site-directed mutagenesis, the mutant shows reduced Vmax and altered Km compared to the wild-type enzyme Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
2'(3')-O-(N-methylanthraniloyl)-ADP slight competitive inhibition Escherichia coli
2'(3')-O-(N-methylanthraniloyl)-AMP slight competitive inhibition Escherichia coli
2'(3')-O-(N-methylanthraniloyl)-ATP competitive Escherichia coli
additional information inhibition of mutant catalytic domains by 2'(3')-O-(N-methylanthraniloyl)-modified nucleotides is reduced compared to the wild-type catalytic domain Cya2-446, overview Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.153
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant Y394A, 10 mM Mg2+ Escherichia coli
0.165
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant W374A, 10 mM Mg2+ Escherichia coli
0.168
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant W249A, 10 mM Mg2+ Escherichia coli
0.201
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant K136A, 10 mM Mg2+ Escherichia coli
0.216
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant E242A, 10 mM Mg2+ Escherichia coli
0.259
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant K253A, 10 mM Mg2+ Escherichia coli
0.27
-
 ATP pH 8.0, 37°C, recombinant catalytic domain, 10 mM Mg2+ Escherichia coli
0.275
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant E185A, 10 mM Mg2+ Escherichia coli
0.284
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant K260A, 10 mM Mg2+ Escherichia coli
0.33
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant S113A, 10 mM Mg2+ Escherichia coli
0.398
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant T189A, 10 mM Mg2+ Escherichia coli
0.46
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant R19A, 10 mM Mg2+ Escherichia coli
0.533
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant S106A, 10 mM Mg2+ Escherichia coli
0.682
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant K332A, 10 mM Mg2+ Escherichia coli
4.7
-
 ATP pH 8.0, 37°C, recombinant catalytic domain mutant S103A, 10 mM Mg2+ Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ the class I enzyme shows a requirement for free metal ions in addition to the MgATP2- complex, operating with a two-metal-ion mechanism in analogy to class II and calss III enzymes. The native enzyme shows very little activity when the concentration of Mg2+ is much lower than that of ATP, and activity rises strongly when the concentration of Mg2+ exceeds that of ATP Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP Escherichia coli
-
 3',5'-cyclic-AMP + diphosphate
-
 ?

Organism

Organism UniProt Comment Textmining
Escherichia coli P00936 class I AC gene
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant catalytic domain Cya2-446 from strain BL21(DE3) by nickel affinity chromatography Escherichia coli

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
 the specific activity of the catalytic domain fragment Cya2-446 and of the holoenzyme is independent of the protein concentration Escherichia coli
0.281
-
 purified recombinant catalytic domain mutant S106A, 10 mM Mg2+ Escherichia coli
0.396
-
 purified recombinant catalytic domain mutant K136A, 10 mM Mg2+ Escherichia coli
0.48
-
 purified recombinant catalytic domain mutant W374A, 10 mM Mg2+ Escherichia coli
0.513
-
 purified recombinant catalytic domain mutant Y394A, 10 mM Mg2+ Escherichia coli
0.526
-
 purified recombinant catalytic domain mutant K253A, 10 mM Mg2+ Escherichia coli
0.614
-
 purified recombinant catalytic domain mutant R19A, 10 mM Mg2+ Escherichia coli
0.665
-
 purified recombinant catalytic domain, 10 mM Mg2+ Escherichia coli
0.752
-
 purified recombinant catalytic domain mutant E242A, 10 mM Mg2+ Escherichia coli
0.778
-
 purified recombinant catalytic domain mutant W249A, 10 mM Mg2+ Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP
-
 Escherichia coli 3',5'-cyclic-AMP + diphosphate
-
 ?
ATP S103 is important for substrate binding docking to the gamma-phosphate group of ATP Escherichia coli 3',5'-cyclic-AMP + diphosphate
-
 ?
additional information structure-function relationship, overview Escherichia coli ?
-
 ?

Subunits

Subunits Comment Organism
More structure-function relationship, overview Escherichia coli

Synonyms

Synonyms Comment Organism
class I adenylate cyclase
-
 Escherichia coli
Cya
-
 Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
45
-
 native holoenzyme Escherichia coli
47
-
 recombinant catalytic domain Cya2?446 Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8.5
-
 native holoenzyme Escherichia coli
8.5 9 recombinant catalytic domain Cya2-446 Escherichia coli

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.05
-
 2'(3')-O-(N-methylanthraniloyl)-ATP pH 8.0, 37°C, versus ATP, wild-type catalytic domain Cya2-446 Escherichia coli