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Literature summary for 1.14.13.69 extracted from

  • McCarl, V.; Somerville, M.V.; Ly, M.A.; Henry, R.; Liew, E.F.; Wilson, N.L.; Holmes, A.J.; Coleman, N.V.
    Heterologous expression of Mycobacterium alkene monooxygenases in Gram-positive and Gram-negative bacterial hosts (2018), Appl. Environ. Microbiol., 84, e00397-18 .
    View publication on PubMedView publication on EuropePMC

Application

Application Comment Organism
synthesis alkene MOs are of interest for their potential roles in industrial biocatalysis, most notably for the stereoselective synthesis of epoxides. Development of high-activity recombinant biocatalysts for alkene oxidation Mycolicibacterium chubuense

Cloned(Commentary)

Cloned (Comment) Organism
gene etnD, recombinant enyme expression of the gene cluster EtnABCD in Pseudomonas putida strain KT2440, cloning the EtnABCD gene cluster into Pseudomonas putida KT2440 yields detectable epoxidation of ethene, which is stimulated by the coexpression of cpn60 chaperonins from either Mycobacterium spp. or Escherichia coli, the complete EtnABCD enzyme cannot be expressed in Escherichia coli strains BL21(DE3)(pLysS) or Rosetta2 although recombinant EtnB and EtnD proteins can be purified individually in soluble form. Strong functional expression of EtnABCD is achieved in Mycobacterium smegmatis mc2155 from pUS116 plasmid, yielding high epoxidation activity. Commercially available codon-optimizing Escherichia coli strains, such as Rosetta2 (Novagen), cannot supply all of the required rare codons. Methods, overview. Gene etnH is not essential for MO activity Mycolicibacterium chubuense
recombinant functional expression of the PmoABCD gene cluster in Mycobacterium smegmatis strain mc2155 from pUS116 plasmid Mycolicibacterium chubuense

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ethene + NADH + H+ + O2 Mycolicibacterium chubuense
-
oxirane + NAD+ + H2O
-
?
ethene + NADH + H+ + O2 Mycolicibacterium chubuense NBB4
-
oxirane + NAD+ + H2O
-
?
propene + NADH + H+ + O2 Mycolicibacterium chubuense
-
1,2-epoxypropane + NAD+ + H2O
-
?
propene + NADH + H+ + O2 Mycolicibacterium chubuense NBB4
-
1,2-epoxypropane + NAD+ + H2O
-
?

Organism

Organism UniProt Comment Textmining
Mycolicibacterium chubuense
-
-
-
Mycolicibacterium chubuense NBB4
-
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1-butene + NADH + H+ + O2
-
Mycolicibacterium chubuense 2-ethyloxirane + NAD+ + H2O
-
?
1-butene + NADH + H+ + O2 best substrate Mycolicibacterium chubuense 2-ethyloxirane + NAD+ + H2O
-
?
1-octene + NADH + H+ + O2
-
Mycolicibacterium chubuense 2-hexyloxirane + NAD+ + H2O
-
?
ethene + NADH + H+ + O2
-
Mycolicibacterium chubuense oxirane + NAD+ + H2O
-
?
ethene + NADH + H+ + O2 high activity Mycolicibacterium chubuense oxirane + NAD+ + H2O
-
?
ethene + NADH + H+ + O2
-
Mycolicibacterium chubuense NBB4 oxirane + NAD+ + H2O
-
?
additional information EtnABCD is specialized for the oxidation of gaseous alkenes (C2 to C4), and its activity is much lower on liquid alkenes (C5 to C8), substrate specificity, overview. Cyclopentene and styrene are poor substrates Mycolicibacterium chubuense ?
-
-
additional information PmoABCD is specialized for the oxidation of gaseous alkenes (C2 to C4), and its activity is much lower on liquid alkenes (C5 to C8), substrate specificity, overview. Cyclopentene is a poor substrate Mycolicibacterium chubuense ?
-
-
additional information EtnABCD is specialized for the oxidation of gaseous alkenes (C2 to C4), and its activity is much lower on liquid alkenes (C5 to C8), substrate specificity, overview. Cyclopentene and styrene are poor substrates Mycolicibacterium chubuense NBB4 ?
-
-
additional information PmoABCD is specialized for the oxidation of gaseous alkenes (C2 to C4), and its activity is much lower on liquid alkenes (C5 to C8), substrate specificity, overview. Cyclopentene is a poor substrate Mycolicibacterium chubuense NBB4 ?
-
-
propene + NADH + H+ + O2
-
Mycolicibacterium chubuense 1,2-epoxypropane + NAD+ + H2O
-
?
propene + NADH + H+ + O2 best substrate Mycolicibacterium chubuense 1,2-epoxypropane + NAD+ + H2O
-
?
propene + NADH + H+ + O2
-
Mycolicibacterium chubuense NBB4 1,2-epoxypropane + NAD+ + H2O
-
?
propene + NADH + H+ + O2 best substrate Mycolicibacterium chubuense NBB4 1,2-epoxypropane + NAD+ + H2O
-
?
styrene + NADH + H+ + O2
-
Mycolicibacterium chubuense ? + NAD+ + H2O
-
?

Subunits

Subunits Comment Organism
heterotetramer 1 * 36000, subunit EtnA, + 1 * 12000 subunit EtnB, + 1 * 59000, subunit EtnC, + 1 * 37000, subunit EtnD, SDS-PAGE Mycolicibacterium chubuense
heterotetramer 1 * 39000, subunit PmoA, + 1 * 12000 subunit PmoB, + 1 * 57000, subunit PmoC, + 1 * 37000, subunit PmoD, SDS-PAGE Mycolicibacterium chubuense
More the alkene MO consists of a small hydroxylase subunit EtnA of 36 kDa, a coupling protein subunit EtnB of12 kDa, a large hydroxylase subunit EtnC of 59 kDa, and a reductase subunit EtnD of 37 kDa Mycolicibacterium chubuense
More the alkene MO consists of a small hydroxylase subunit PmoA of 39 kDa, a coupling protein subunit PmoB of12 kDa, a large hydroxylase subunit PmoC of 57 kDa, and a reductase subunit PmoD of 37 kDa Mycolicibacterium chubuense

Synonyms

Synonyms Comment Organism
ethene MO
-
Mycolicibacterium chubuense
EtnABCD
-
Mycolicibacterium chubuense
etnD
-
Mycolicibacterium chubuense
PmoABCD
-
Mycolicibacterium chubuense
propene MO
-
Mycolicibacterium chubuense

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
assay at Mycolicibacterium chubuense

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
assay at Mycolicibacterium chubuense

Cofactor

Cofactor Comment Organism Structure
FAD
-
Mycolicibacterium chubuense
NADH
-
Mycolicibacterium chubuense

General Information

General Information Comment Organism
evolution the enzyme belongs to the bacterial monooxygenases of the soluble di-iron MOs (SDIMOs) type, thereof the group 4 SDIMOs are four-component alkene monooxygenases from bacteria that grow on ethene and/or propene. Despite their sequence and substrate diversity, all the SDIMOs have similar biochemistries. Electrons are transferred from NADH to an oxidoreductase protein that contains flavin and iron-sulfur clusters, and thence to a catalytic hydroxylase (made of 2 to 3 proteins) that contains the binuclear iron active site. In the active site, one oxygen atom in O2 is reduced to water, while the other is activated to a high-energy state and attacks the substrate. The catalytic activity is acilitated by a small cofactor-independent coupling protein, and depending on the SDIMO family, other proteins, such as ferredoxins, may also be part of the MO-enzyme complex. Mycobacterium chubuense NBB4 is unique among hydrocarbon-oxidizing bacteria because it contains four different SDIMOs, in addition to a copper-containing MO, a P450, and an alkB homologue. Strain NBB4 has two group 4 SDIMOs (etnABCD and pmoABCD), an atypical group 3 SDIMO (smoXYB1C1Z), and a group 6 SDIMO (smoABCD). To date, there is experimental evidence that smoXYB1C1Z is a gaseous alkane/alkene MO and that etnABCD is an ethene MO. Comparison of SDIMO activities in whole cells of wild-type and recombinant bacteria, overview Mycolicibacterium chubuense