Literature summary for 1.14.13.33 extracted from

Westphal, A.H.; Tischler, D.; Heinke, F.; Hofmann, S.; Groening, J.A.D.; Labudde, D.; van Berkel, W.J.H.
Pyridine nucleotide coenzyme specificity of p-hydroxybenzoate hydroxylase and related flavoprotein monooxygenases (2018), Front. Microbiol., 9, 3050 .

Search for more literature for 1.14.13.33

Cloned(Commentary)

Cloned (Comment)	Organism
gene pobA, DNA and amino acid sequence determination and analysis and tree, sequence comparisons and phylogenetic analysis, recombinant expression of optimized enzyme, subcloning in Escherichia coli strain DH5alpha	Rhodococcus opacus
gene pobA, DNA and amino acid sequence determination and analysis and tree, sequence comparisons and phylogenetic analysis, recombinant expression of optimized enzyme, subcloning in Escherichia coli strain DH5alpha	Rhodococcus rhodnii
gene Reut_B5020, DNA sequence comparisons and phylogenetic analysis and tree, recombinant expression of the codon-optimized gene in Acinetobacter sp. ADP1	Cupriavidus pinatubonensis

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.0194	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADPH	Cupriavidus pinatubonensis
0.0202	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADH	Cupriavidus pinatubonensis
0.0204	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADPH	Cupriavidus pinatubonensis
0.0304	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
0.035	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination, with NADPH	Rhodococcus opacus
0.0397	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
0.042	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC, with NADPH	Rhodococcus opacus
0.0498	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination	Cupriavidus pinatubonensis
0.0499	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC	Rhodococcus opacus
0.146	-	NADPH	pH 7.5, 30°C, recombinant enzyme PHBHCn1, UV/VIS-NAD(P)H consumption determination	Cupriavidus pinatubonensis
0.153	-	NADPH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
0.193	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADH	Cupriavidus pinatubonensis

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
4-hydroxybenzoate + NADH + H+ + O2	Cupriavidus pinatubonensis	-	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	Rhodococcus rhodnii	-	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	Rhodococcus opacus	preferred substrates	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	Rhodococcus rhodnii 135	-	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	Rhodococcus opacus 557	preferred substrates	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	Cupriavidus pinatubonensis	-	3,4-dihydroxybenzoate + NADP+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	Rhodococcus opacus	-	3,4-dihydroxybenzoate + NADP+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	Rhodococcus opacus 557	-	3,4-dihydroxybenzoate + NADP+ + H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Cupriavidus pinatubonensis	Q46R66	Cupriavidus necator or Ralstonia eutropha	-
Rhodococcus opacus	A0A0U1URQ7	-	-
Rhodococcus opacus 557	A0A0U1URQ7	-	-
Rhodococcus rhodnii	A0A0U1URV4	-	-
Rhodococcus rhodnii 135	A0A0U1URV4	-	-

Source Tissue

Source Tissue	Comment	Organism	Textmining
additional information	culture growth with 4-hydroxybenzoate as sole source of carbon and energy	Rhodococcus opacus	-
additional information	culture growth with 4-hydroxybenzoate as sole source of carbon and energy	Rhodococcus rhodnii	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
4-hydroxybenzoate + NADH + H+ + O2	-	Cupriavidus pinatubonensis	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	-	Rhodococcus rhodnii	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	preferred substrates	Rhodococcus opacus	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	-	Rhodococcus rhodnii 135	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADH + H+ + O2	preferred substrates	Rhodococcus opacus 557	3,4-dihydroxybenzoate + NAD+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	-	Cupriavidus pinatubonensis	3,4-dihydroxybenzoate + NADP+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	-	Rhodococcus opacus	3,4-dihydroxybenzoate + NADP+ + H2O	-	?
4-hydroxybenzoate + NADPH + H+ + O2	-	Rhodococcus opacus 557	3,4-dihydroxybenzoate + NADP+ + H2O	-	?
additional information	enzyme PHBHRo shows a clear preference for NADH compared to NADPH	Rhodococcus opacus	?	-	-
additional information	enzyme PHBHRr shows a clear preference for NADH compared to NADPH	Rhodococcus rhodnii	?	-	-
additional information	enzyme PHBHRr shows a clear preference for NADH compared to NADPH	Rhodococcus rhodnii 135	?	-	-
additional information	enzyme PHBHRo shows a clear preference for NADH compared to NADPH	Rhodococcus opacus 557	?	-	-

Subunits

Subunits	Comment	Organism
More	PHBH and related enzymes lack a canonical NAD(P)H-binding domain	Cupriavidus pinatubonensis
More	PHBH and related enzymes lack a canonical NAD(P)H-binding domain	Rhodococcus opacus
More	PHBH and related enzymes lack a canonical NAD(P)H-binding domain	Rhodococcus rhodnii

Synonyms

Synonyms	Comment	Organism
NAD(P)H-dependent PHBHCn1	-	Cupriavidus pinatubonensis
NADPH-dependent PHBH	UniProt	Rhodococcus opacus
p-hydroxybenzoate hydroxylase	-	Cupriavidus pinatubonensis
p-hydroxybenzoate hydroxylase	-	Rhodococcus opacus
p-hydroxybenzoate hydroxylase	-	Rhodococcus rhodnii
PHBH	-	Cupriavidus pinatubonensis
PHBH	-	Rhodococcus opacus
PHBH	-	Rhodococcus rhodnii
PHBHCn1	-	Cupriavidus pinatubonensis
PHBHRo	-	Rhodococcus opacus
PHBHRo1CP	-	Rhodococcus opacus
PHBHRr	-	Rhodococcus rhodnii
PobA	-	Rhodococcus opacus
PobA	-	Rhodococcus rhodnii
Reut_B5020	-	Cupriavidus pinatubonensis

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Cupriavidus pinatubonensis
30	-	assay at	Rhodococcus opacus
30	-	assay at	Rhodococcus rhodnii

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
4.6	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADPH	Cupriavidus pinatubonensis
6.5	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADPH	Cupriavidus pinatubonensis
6.8	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADH	Cupriavidus pinatubonensis
7.6	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADH	Cupriavidus pinatubonensis
8.7	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination, with NADPH	Rhodococcus opacus
9.4	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination	Cupriavidus pinatubonensis
9.6	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC, with NADPH	Rhodococcus opacus
12.9	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination, with NADH	Rhodococcus opacus
15	-	NADPH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
15.3	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
16.8	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC, with NADH	Rhodococcus opacus

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Cupriavidus pinatubonensis
7.5	-	assay at	Rhodococcus opacus
7.5	-	assay at	Rhodococcus rhodnii

Cofactor

Cofactor	Comment	Organism	Structure
FAD	the enzyme is a flavoprotein	Cupriavidus pinatubonensis
FAD	the enzyme is a flavoprotein	Rhodococcus opacus
FAD	the enzyme is a flavoprotein	Rhodococcus rhodnii
NADH	PHBH and related enzymes lack a canonical NAD(P)H-binding domain enzyme PHBHRr shows a clear preference for NADH	Rhodococcus rhodnii
NADH	PHBH and related enzymes lack a canonical NAD(P)H-binding domain. Enzyme PHBHRo shows a clear preference for NADH, PHBHRo1CP contains the NADH-preferring sequence motif 32-ESRTREEVEGT. NADH is preferred over NADPHs	Rhodococcus opacus
NADH	PHBH and related enzymes lack a canonical NAD(P)H-binding domain. NAD(P)H-dependent PHBHCn1	Cupriavidus pinatubonensis
NADPH	NADH is preferred before NADPH	Rhodococcus opacus
NADPH	PHBH and related enzymes lack a canonical NAD(P)H-binding domain. NAD(P)H-dependent PHBHCn1	Cupriavidus pinatubonensis

General Information

General Information	Comment	Organism
evolution	amino acid sequences of NADH-preferring PHBHs of putative PHBHs identified in currently available bacterial genomes, phylogenetic analysis, overview. The pyridine nucleotide coenzyme specificity of PHBH emerged through adaptive evolution, and the NADH-preferring enzymes are the older versions of PHBH. Structural comparison and distance tree analysis of group A flavoprotein monooxygenases indicates that a similar protein segment as being responsible for the pyridine nucleotide coenzyme specificity of PHBH is involved in determining the pyridine nucleotide coenzyme specificity of the other group A members. Evolutionary rate calculation. Among the actinobacterial sequences presently available, most comprise the NADH-preferring fingerprint. However, Mycobacteria have a mixed type motif, often the first or both arginine(s) of the NADH-fingerprint are present but the remaining part is lacking. In addition, many mycobacterial sequences have parts of the NADPH-preferring fingerprint, especially, x(D/E)YVL(G/S)R	Cupriavidus pinatubonensis
evolution	amino acid sequences of NADH-preferring PHBHs of putative PHBHs identified in currently available bacterial genomes, phylogenetic analysis, overview. The pyridine nucleotide coenzyme specificity of PHBH emerged through adaptive evolution, and the NADH-preferring enzymes are the older versions of PHBH. Structural comparison and distance tree analysis of group A flavoprotein monooxygenases indicates that a similar protein segment as being responsible for the pyridine nucleotide coenzyme specificity of PHBH is involved in determining the pyridine nucleotide coenzyme specificity of the other group A members. Evolutionary rate calculation. Among the actinobacterial sequences presently available, most comprise the NADH-preferring fingerprint. However, Mycobacteria have a mixed type motif, often the first or both arginine(s) of the NADH-fingerprint are present but the remaining part is lacking. In addition, many mycobacterial sequences have parts of the NADPH-preferring fingerprint, especially, x(D/E)YVL(G/S)R	Rhodococcus opacus
evolution	amino acid sequences of NADH-preferring PHBHs of putative PHBHs identified in currently available bacterial genomes, phylogenetic analysis, overview. The pyridine nucleotide coenzyme specificity of PHBH emerged through adaptive evolution, and the NADH-preferring enzymes are the older versions of PHBH. Structural comparison and distance tree analysis of group A flavoprotein monooxygenases indicates that a similar protein segment as being responsible for the pyridine nucleotide coenzyme specificity of PHBH is involved in determining the pyridine nucleotide coenzyme specificity of the other group A members. Evolutionary rate calculation. Among the actinobacterial sequences presently available, most comprise the NADH-preferring fingerprint. However, Mycobacteria have a mixed type motif, often the first or both arginine(s) of the NADH-fingerprint are present but the remaining part is lacking. In addition, many mycobacterial sequences have parts of the NADPH-preferring fingerprint, especially, x(D/E)YVL(G/S)R	Rhodococcus rhodnii
additional information	energy profiling from enzyme protein structure is realized by means of a coarse-grained residue-level pair potential function modeling, overview	Cupriavidus pinatubonensis
additional information	energy profiling from enzyme protein structure is realized by means of a coarse-grained residue-level pair potential function modeling, overview	Rhodococcus opacus
additional information	energy profiling from enzyme protein structure is realized by means of a coarse-grained residue-level pair potential function modeling, overview	Rhodococcus rhodnii

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
73.1	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADH	Cupriavidus pinatubonensis
98.04	-	NADPH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
188.8	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination	Cupriavidus pinatubonensis
225.5	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via product determination per HPLC, with NADPH	Cupriavidus pinatubonensis
228.6	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC, with NADPH	Rhodococcus opacus
248.6	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination, with NADPH	Rhodococcus opacus
335.1	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADPH	Cupriavidus pinatubonensis
336.7	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via product determination per HPLC	Rhodococcus opacus
376.2	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHCn1, via UV/VIS-NAD(P)H consumption determination, with NADH	Cupriavidus pinatubonensis
385.4	-	NADH	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus
424.3	-	4-hydroxybenzoate	pH 7.5, 30°C, recombinant enzyme PHBHRo1CP, via UV/VIS-NAD(P)H consumption determination	Rhodococcus opacus

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Release 2023.1 (January 2023)