Pynr071c-α factor-curA-AOX1 Terminator
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21Illegal XhoI site found at 1438
- 23COMPATIBLE WITH RFC
- 25COMPATIBLE WITH RFC
- 1000COMPATIBLE WITH RFC
This is a composite part for extracellular expression of NADPH-dependent curcumin.curA is expressed and participates in the production from curcumin to curcumin reductase. Pynr071c is the promoter of Aldose 1-isomerase superfamily protein YNR071C. In the presence of xylose, the signal peptide, α-factor , is used to express curA outside of the cell.
Usage and Biology
curA is derived from curcumin-transforming microorganisms isolated from human faeces with a molecular weight of about 82 kDa and consists of two identical subunits. During the purification of the expressed curA enzyme, the activity of curA enzyme was lost during dialysis. However, we found that the addition of NADPH increased the activity, but the addition of NADH did not increase the activity. These findings indicate that the enzyme catalyzes the NADPH-dependent transformation of curcumin. In the curA catalytic reaction, two steps of curcumin metabolism pathway (curcumin→dihydrocurcumin→tetrahydrocurcumin) were found. Under the catalysis of CurA, dihydrocurcumin is first the product, then the substrate. These products are produced from curcumin by reducing the diarylheptatrienone chain which could devastate the color of the hair. The curA does not produce more reduction products than tetrahydrocurcumin, which indicates that curA only catalyzes the reduction of compounds with C=C. In the reaction process, the optimum reaction temperature of curA is 35 ℃. The enzyme showed maximum activity at pH 5.9. Although a large number of different compounds have been tested as potential substrates of curA, curA seems to have a narrow substrate spectrum and preferentially acts on curcumin. Considering substrate specificity and NADPH dependence, curA was named NADPH-dependent curcumin/dihydrocurcumin reductase.