Literature summary for 1.14.15.1 extracted from

Bell, S.G.; Yang, W.; Dale, A.; Zhou, W.; Wong, L.L.
Improving the affinity and activity of CYP101D2 for hydrophobic substrates (2013), Appl. Microbiol. Biotechnol., 97, 3979-3990.

Search for more literature for 1.14.15.1

Cloned(Commentary)

Cloned (Comment)	Organism
expression of N-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)	Novosphingobium aromaticivorans

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant His-tagged wild-type and mutant enzymes, hanging drop vapour diffusion method, mixing of 0.001 ml of 50 mg/ml protein in crystallisation buffer containing 20 mM Tris, pH 8.0, and 150 mM KCl, with 0.001 ml of reservoir solution containing 0.1 M Tris, pH 8.3, 5% v/v PEG 400, and 1.9 M ammonium sulfate, equilivration against 0.1 ml of reservoir solution, 18°C, X-ray diffraction structure determination and analysis at 2.0 A resolution	Novosphingobium aromaticivorans

Crystallization (Comment)

Organism

purified recombinant His-tagged wild-type and mutant enzymes, hanging drop vapour diffusion method, mixing of 0.001 ml of 50 mg/ml protein in crystallisation buffer containing 20 mM Tris, pH 8.0, and 150 mM KCl, with 0.001 ml of reservoir solution containing 0.1 M Tris, pH 8.3, 5% v/v PEG 400, and 1.9 M ammonium sulfate, equilivration against 0.1 ml of reservoir solution, 18°C, X-ray diffraction structure determination and analysis at 2.0 A resolution

Novosphingobium aromaticivorans

Protein Variants

Protein Variants	Comment	Organism
L253V	site-directed mutagenesis, the mutant shows 95% reduced activity compared to the wild-type enzyme, crystal structure analysis of mutant enzyme with bound substrate	Novosphingobium aromaticivorans
M98F	site-directed mutagensis, the mutant shows 85% reduced activity compared to the wild-type enzyme, crystal structure analysis of mutant enzyme with bound substrate	Novosphingobium aromaticivorans
additional information	substitutions at Tyr96, Met98 and Leu253 in CYP101D2 analogously to closely related CYP101A1 from Pseudomonas putida, reduce both the spin state shift on camphor binding and the camphor oxidation activity	Novosphingobium aromaticivorans
Y96A	site-directed mutagensis, the mutant shows increased affinity for hydrocarbon substrates including adamantane, cyclooctane, hexane and 2-methylpentane, the monooxygenase activity of the mutant towards alkane substrates is enhanced compared to the wild-type enzyme, crystal structure analysis of mutant enzyme with bound substrate	Novosphingobium aromaticivorans

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Fe2+	heme and cytochrome P450 containing enzyme	Novosphingobium aromaticivorans

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
(+)-camphor + reduced ferredoxin + O2	Novosphingobium aromaticivorans	-	(+)-exo-5-hydroxycamphor + oxidized ferredoxin + H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Novosphingobium aromaticivorans	Q2G8A2	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N--terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3)	Novosphingobium aromaticivorans

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
(+)-camphor + reduced ferredoxin + O2	-	Novosphingobium aromaticivorans	(+)-exo-5-hydroxycamphor + oxidized ferredoxin + H2O	-	?
(+)-camphor + reduced ferredoxin + O2	-	Novosphingobium aromaticivorans	(+)-exo-5-hydroxycamphor + oxidized ferredoxin + H2O	99% 5-exo-hydroxycamphor by wild-type enzyme and 92% by Y96A mutant	?
3 2-methylpentane + 3 reduced ferreredoxin + O2	-	Novosphingobium aromaticivorans	2-methyl-pentan-2-ol + 2-methyl-pentan-3-ol + 2-methyl-pentan-4-ol + 3 oxidized ferredoxin	52.5% 2-methyl-pentan-2-ol + 13% 2-methyl-pentan-3-ol, and 3% 2-methyl-pentan-4-ol for the wild-type enzyme, 5% + 12% + 30% for the mutant Y96A	?
adamantane + reduced ferreredoxin + O2	-	Novosphingobium aromaticivorans	1-adamantol + 2-adamantol + oxidized ferredoxin + H2O	98% 1-adamantol + 2% 2-adamantol for the wild-type enzyme, 97% + 3% for the mutant Y96A	?
cyclooctane + reduced ferreredoxin + O2	-	Novosphingobium aromaticivorans	cyclooctanol + cyclooctanone + oxidized ferredoxin + H2O	99% cyclooctanol + 1% cyclooctanone for the wild-type enzyme, 97% + 3% for the mutant Y96A	?
hexane + reduced ferreredoxin + O2	-	Novosphingobium aromaticivorans	hexan-2-ol + hexan-3-ol + oxidized ferredoxin + H2O	56% hexan-2-ol + 44% hexan-3-ol for the wild-type enzyme and mutant Y96A	?
additional information	substrate binding and activity data for wild-type CYP101D2 and variants with different substrates, gas chromatography analysis of products, overview	Novosphingobium aromaticivorans	?	-	?

Synonyms

Synonyms	Comment	Organism
CYP101D2	-	Novosphingobium aromaticivorans

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Novosphingobium aromaticivorans

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.4	-	assay at	Novosphingobium aromaticivorans

Cofactor

Cofactor	Comment	Organism
cytochrome P450	CYP101D2 is a cytochrome P450 monooxygenase	Novosphingobium aromaticivorans
Ferredoxin	[2Fe-2S] ferredoxin	Novosphingobium aromaticivorans
heme	heme-dependent monooxygenase	Novosphingobium aromaticivorans

General Information

General Information	Comment	Organism
additional information	role of Tyr96 in substrate binding	Novosphingobium aromaticivorans