Cloned (Comment) | Organism |
---|---|
gene paaK1, paaK1 is a single copy gene, recombinant expression of His-tagged isozyme PaaK1 in Escherichia coli strain BL21Star(DE3), expression of selenomethionine-labeled PaaK1 in Escherichia coli strain 834(DE3) | Burkholderia cenocepacia |
gene paaK2, recombinant expression of His-tagged isozyme PaaK2 in Escherichia coli strain BL21Star(DE3) | Burkholderia cenocepacia |
Crystallization (Comment) | Organism |
---|---|
adenylated phenylacetate intermediate complex of purified recombinant His-tagged PaaK2, sitting drop vapor diffusion method, 12 mg/ml protein in 20 mM HEPES, pH 8.3, 10 mM NaCl, 3 mM 2-mercaptoethanol, 2% glycerol, 3 mM MgCl2, and 3 mM ATP, is mixed with 17% w/v PEG 6000, 0.1 M HEPES, pH 7.5, 0.1 M KCl, and 2.5% glycerol, X-ray diffraction structure determination and analysis at 1.90 A resolution | Burkholderia cenocepacia |
purified recombinant His-tagged selenomethionine derivative of PaaK1 in complex with ATP and adenylated phenylacetate intermediate complex of purified recombinant His-tagged PaaK1, sitting drop vapor diffusion method, 12 mg/ml protein in 20 mM HEPES, pH 8.3, 10 mM NaCl, 3 mM 2-mercaptoethanol, 2% glycerol, 3 mM MgCl2, and 3 mM ATP, is mixed with 20-25% PEG 3350, 200 mM potassium thiocyanate, and 5% glycerol. Crystals of PaaK1 are grown from 10% w/v PEG 8000, 0.1 M HEPES, pH 7.5, and 8.0% v/v ethylene glycol, X-ray diffraction structure determination and analysis at 1.6 A and 1.92 A resolution, respectively | Burkholderia cenocepacia |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | isozyme PaaK1 possesses a lower Km for phenylacetic acid and is better able to accommodate 3' and 4' substitutions on the phenyl ring than isozyme PaaK2 | Burkholderia cenocepacia | |
0.062 | - |
phenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
0.15 | - |
phenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
0.17 | - |
3-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
0.23 | - |
3-Chlorophenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
0.54 | - |
4-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
0.87 | - |
3-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
1.5 | - |
3-Chlorophenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required, the beta- and gamma-phosphates of ATP coordinate a single Mg2+ ion that is further coordinated by four water molecules in an overall octahedral geometry | Burkholderia cenocepacia |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + phenylacetate + CoA | Burkholderia cenocepacia | - |
AMP + diphosphate + phenylacetyl-CoA | - |
? | |
ATP + phenylacetate + CoA | Burkholderia cenocepacia DSM 16553 | - |
AMP + diphosphate + phenylacetyl-CoA | - |
? | |
additional information | Burkholderia cenocepacia | isozyme PaaK1 displays a lower Km for phenylacetic acid and broader substrate specificity than isozyme PaaK2 | ? | - |
? | |
additional information | Burkholderia cenocepacia DSM 16553 | isozyme PaaK1 displays a lower Km for phenylacetic acid and broader substrate specificity than isozyme PaaK2 | ? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Burkholderia cenocepacia | B4E7B5 | gene paaK1 | - |
Burkholderia cenocepacia | B4EL89 | gene paaK2 | - |
Burkholderia cenocepacia DSM 16553 | B4E7B5 | gene paaK1 | - |
Burkholderia cenocepacia DSM 16553 | B4EL89 | gene paaK2 | - |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged isozyme PaaK1 from Escherichia coli strain BL21Star(DE3) and selenomethionine-labeled His-tagged PaaK1 from Escherichia coli strain 834(DE3) by nickel affinity chromatography, gel filtration, and anion exchange chromatography | Burkholderia cenocepacia |
recombinant His-tagged isozyme PaaK2 from Escherichia coli strain BL21Star(DE3) by nickel affinity chromatography, gel filtration, and anion exchange chromatography | Burkholderia cenocepacia |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + 3-chlorophenylacetate + CoA | - |
Burkholderia cenocepacia | AMP + diphosphate + 3-chlorophenylacetyl-CoA | - |
? | |
ATP + 3-chlorophenylacetate + CoA | - |
Burkholderia cenocepacia DSM 16553 | AMP + diphosphate + 3-chlorophenylacetyl-CoA | - |
? | |
ATP + 3-hydroxyphenylacetate + CoA | - |
Burkholderia cenocepacia | AMP + diphosphate + 3-hydroxyphenylacetyl-CoA | - |
? | |
ATP + 3-hydroxyphenylacetate + CoA | - |
Burkholderia cenocepacia DSM 16553 | AMP + diphosphate + 3-hydroxyphenylacetyl-CoA | - |
? | |
ATP + 4-hydroxyphenylacetate + CoA | - |
Burkholderia cenocepacia | AMP + diphosphate + 4-hydroxyphenylacetyl-CoA | - |
? | |
ATP + 4-hydroxyphenylacetate + CoA | - |
Burkholderia cenocepacia DSM 16553 | AMP + diphosphate + 4-hydroxyphenylacetyl-CoA | - |
? | |
ATP + phenylacetate + CoA | - |
Burkholderia cenocepacia | AMP + diphosphate + phenylacetyl-CoA | - |
? | |
ATP + phenylacetate + CoA | - |
Burkholderia cenocepacia DSM 16553 | AMP + diphosphate + phenylacetyl-CoA | - |
? | |
additional information | isozyme PaaK1 displays a lower Km for phenylacetic acid and broader substrate specificity than isozyme PaaK2 | Burkholderia cenocepacia | ? | - |
? | |
additional information | no activity 4-hydroxyphenylacetate, 3,4-dihydroxyphenylacetate and 2-chloro-2-hydroxy-phenylacetate | Burkholderia cenocepacia | ? | - |
? | |
additional information | no activity with 3,4-dihydroxyphenylacetate and 2-chloro-2-hydroxy-phenylacetate | Burkholderia cenocepacia | ? | - |
? | |
additional information | isozyme PaaK1 displays a lower Km for phenylacetic acid and broader substrate specificity than isozyme PaaK2 | Burkholderia cenocepacia DSM 16553 | ? | - |
? | |
additional information | no activity with 3,4-dihydroxyphenylacetate and 2-chloro-2-hydroxy-phenylacetate | Burkholderia cenocepacia DSM 16553 | ? | - |
? | |
additional information | no activity 4-hydroxyphenylacetate, 3,4-dihydroxyphenylacetate and 2-chloro-2-hydroxy-phenylacetate | Burkholderia cenocepacia DSM 16553 | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
PaaK1 | - |
Burkholderia cenocepacia |
PaaK2 | - |
Burkholderia cenocepacia |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
1.17 | - |
4-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
1.6 | - |
3-Chlorophenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
1.67 | - |
3-Chlorophenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
1.83 | - |
3-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
2.67 | - |
3-hydroxyphenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
4.17 | - |
phenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia | |
5 | - |
phenylacetate | pH and temperature not specified in the publication | Burkholderia cenocepacia |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
ATP | - |
Burkholderia cenocepacia |
General Information | Comment | Organism |
---|---|---|
evolution | PaaK1 and PaaK2 form a unique subgroup within the adenylate-forming enzyme superfamily | Burkholderia cenocepacia |
additional information | adenylated phenylacetate intermediate complexes of PaaK1 and PaaK2 occur in distinct conformations, a N-terminal microdomain may serve to recruit subsequent phenylacetate enzymes, whereas a bifunctional role is proposed for the P-loop in stabilizing the C-terminal domain in conformation 2, an extended aryl binding pocket in PaaK1 contrasts with PaaK2, overview | Burkholderia cenocepacia |
additional information | isozyme PaaK1 shows dynamic enzyme-substrate interactions. Adenylated phenylacetate intermediate complexes of PaaK1 and PaaK2 occur in distinct conformations, a N-terminal microdomain may serve to recruit subsequent phenylacetate enzymes, whereas a bifunctional role is proposed for the P-loop in stabilizing the C-terminal domain in conformation 2, an extended aryl binding pocket in PaaK1 contrasts with PaaK2, overview | Burkholderia cenocepacia |
physiological function | the phenylacetic acid degradation pathway is the sole aerobic route for phenylacetic acid metabolism in bacteria and facilitates degradation of environmental pollutants such as styrene and ethylbenzene. The PAA pathway also is implicated in promoting Burkholderia cenocepacia infections in cystic fibrosis patients | Burkholderia cenocepacia |
physiological function | the phenylacetic acid degradation pathway is the sole aerobic route for phenylacetic acid metabolism in bacteria and facilitates degradation of environmental pollutants such as styrene and ethylbenzene. The phenylacetic acid pathway also is implicated in promoting Burkholderia cenocepacia infections in cystic fibrosis patients | Burkholderia cenocepacia |