RRC ID |
55835
|
著者 |
Tamaki S, Yagi M, Nishihata Y, Yamaji H, Shigeri Y, Uno T, Imaishi H.
|
タイトル |
Modification of N-Terminal Amino Acids of Fungal Benzoate Hydroxylase (CYP53A15) for the Production of p-Hydroxybenzoate and Optimization of Bioproduction Conditions in Escherichia coli.
|
ジャーナル |
J Microbiol Biotechnol
|
Abstract |
The aromatic compound p-hydroxybenzoate (PHBA) is an important material with multiple applications, including as a building block of liquid crystal polymers in chemical industries. The cytochrome P450 (CYP) enzymes are beneficial monooxygenases for the synthesis of chemicals, and CYP53A15 from fungus Cochliobolus lunatus is capable of executing the hydroxylation from benzoate to PHBA. Here, we constructed a system for the bioconversion of benzoate to PHBA in Escherichia coli cells coexpressing CYP53A15 and human NADPH-P450 oxidoreductase (CPR) genes as a redox partner. For suitable coexpression of CYP53A15 and CPR, we originally constructed five plasmids in which we replaced the N-terminal transmembrane region of CYP53A15 with a portion of the N-terminus of various mammalian P450s. PHBA productivity was the greatest when CYP53A15 expression was induced at 20°C in 2×YT medium in host E. coli strain ΔgcvR transformed with an N-terminal transmembrane region of rabbit CYP2C3. By optimizing each reaction condition (reaction temperature, substrate concentration, reaction time, and E. coli cell concentration), we achieved 90% whole-cell conversion of benzoate. Our data demonstrate that the described novel E. coli bioconversion system is a more efficient tool for PHBA production from benzoate than the previously described yeast system.
|
巻・号 |
28(3)
|
ページ |
439-447
|
公開日 |
2018-3-28
|
DOI |
10.4014/jmb.1711.11030
|
PII |
10.4014/jmb.1711.11030
|
PMID |
29539883
|
MeSH |
Animals
Ascomycota / enzymology*
Ascomycota / genetics*
Benzoates / metabolism
Cloning, Molecular
Cytochrome P-450 Enzyme System / metabolism
Escherichia coli / genetics*
Escherichia coli / metabolism*
Gene Expression Regulation, Bacterial
Genes, Fungal / genetics
Humans
NADPH-Ferrihemoprotein Reductase / genetics
Oxygenases / chemistry*
Oxygenases / genetics*
Parabens / metabolism*
Plasmids
Rabbits
Reaction Time
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Temperature
|
IF |
1.65
|
引用数 |
1
|
リソース情報 |
原核生物(大腸菌) |
BW25113, ΔgcvR, ΔcpxA, ΔglnL |