An enone reductase from Nicotiana tabacum: cDNA cloning, expression in Escherichia coli, and reduction of enones with the recombinant proteins.
In the course of the purification of enone reductase participating to the reduction of pulegone, two reductases (NtRed-1 and NtRed-2) were isolated from cultured cells of Nicotiana tabacum. The partial amino acid sequences of the reductases revealed that NtRed-1 was allyl-alcohol dehydrogenase (Accession No. BAA89423) and NtRed-2 was malate dehydrogenase (Accession No. CAC12826). cDNA cloning and expression of these reductases in Escherichia coli were performed. Reduction with recombinant proteins was examined with cyclic alpha,beta-unsaturated ketones, such as pulegone, carvone and verbenone, as substrates. It was found that the recombinant NtRed-1 catalyses the hydrogenation of the exocyclic C-C double bond of pulegone.
Matsushima
Akihito
A
Natural Science Center for Basic Research and Development, Radioisotope Center, Hiroshima University, 1-4-2 Kagamiyama, Higashi-Hiroshima 739-8526, Japan.
Sato
Yuya
Y
Otsuka
Miki
M
Watanabe
Takayoshi
T
Yamamoto
Hiroaki
H
Hirata
Toshifumi
T
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D007659:Ketones / Q000737:chemistry / Q000378:metabolism*; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014026:Nicotiana / Q000201:enzymology*
D018076:DNA, Complementary; D007659:Ketones; D011994:Recombinant Proteins; D010088:Oxidoreductases