Part:BBa_K4825059
pTEF2-CDC9-LEU1-tSSA1-pPGK1-COX6-LEU2-tPGK1-pTDH3-COX4-LEU4^S547∆-tTDH1
CDC9 gene from yeast encodes the activity required during DNA replication by DNA ligase I. The N- terminal localization signals from CDC9 was used to taget Leu1p in the mitochondria, the first 47 amino acids from CDC9 were appended to the N- terminus of the open reading frame. LEU1 is an endogenous gene in yeast 2-ketoacid elongation pathway. It encodes for isopropylmalate Isomerase, which isomerizes 2-isopropylmalate(2-IPM), an intermediate in the pathway, to produce 3-isopropylmalate(3-IPM). isopropylmalate Isomerase naturally functions in cytosol.
LEU2, encoding IPM dehydrogenase, is an endogenous gene in yeast 2-ketoacid elongation pathway. The enzyme oxidizes and decarboxylates 3 IPM to produce 2-ketoisocaproate(KIC), and important precursor for isoamyl acetate. IPM dehydrogenase naturally function in cytosol.
LEU4 encodes for 2-isopropylmalate synthase(2-IPMS), which acetylates KIV and turning it into 2-isopropylmalate(2-IPM). This is one of the three reaction in 2-ketoacid elongation pathway. 2-IPMS naturally locates both in mitochondria and cytosol.
The terminal enzyme in the mitochondrial respiratory chain is COX. COX contains 13 mitochondrial and nuclear encoded subunits. The nuclear encoded subunits have yet unknown function but are hypothesized to play a role in constructing and regulating the complex. The mitochondrial encoded subunits aids in the performance of the ETC and proton pumping activities. The N- terminal localization signals from COXIV is used to target Leu4p to mitochondria, and the first 26 amino acids from COX4 appends to the N-terminus of the corresponding open reading frame.
COX6 is the last enzyme in line in the mitochondrial electron transport chain, driving oxidative phosphorylation. it is a component of the cytochrome c oxidase. The N- terminal localization signals from COXVI were used to target Leu2p to mitochondria, and the first 41 amino acids from the COX VI were appended to the N- terminus of the corresponding open reading frame.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 1572
Illegal EcoRI site found at 2036
Illegal EcoRI site found at 3326
Illegal EcoRI site found at 4115
Illegal EcoRI site found at 6579
Illegal EcoRI site found at 7518
Illegal XbaI site found at 1813
Illegal XbaI site found at 8965
Illegal XbaI site found at 9775
Illegal XbaI site found at 9889
Illegal SpeI site found at 9930
Illegal PstI site found at 2502
Illegal PstI site found at 3300
Illegal PstI site found at 3795
Illegal PstI site found at 4080 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1572
Illegal EcoRI site found at 2036
Illegal EcoRI site found at 3326
Illegal EcoRI site found at 4115
Illegal EcoRI site found at 6579
Illegal EcoRI site found at 7518
Illegal NheI site found at 9453
Illegal SpeI site found at 9930
Illegal PstI site found at 2502
Illegal PstI site found at 3300
Illegal PstI site found at 3795
Illegal PstI site found at 4080 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1572
Illegal EcoRI site found at 2036
Illegal EcoRI site found at 3326
Illegal EcoRI site found at 4115
Illegal EcoRI site found at 6579
Illegal EcoRI site found at 7518
Illegal BglII site found at 6405
Illegal BglII site found at 10660
Illegal BamHI site found at 9606
Illegal BamHI site found at 10572
Illegal XhoI site found at 5048
Illegal XhoI site found at 11251 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 1572
Illegal EcoRI site found at 2036
Illegal EcoRI site found at 3326
Illegal EcoRI site found at 4115
Illegal EcoRI site found at 6579
Illegal EcoRI site found at 7518
Illegal XbaI site found at 1813
Illegal XbaI site found at 8965
Illegal XbaI site found at 9775
Illegal XbaI site found at 9889
Illegal SpeI site found at 9930
Illegal PstI site found at 2502
Illegal PstI site found at 3300
Illegal PstI site found at 3795
Illegal PstI site found at 4080 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 1572
Illegal EcoRI site found at 2036
Illegal EcoRI site found at 3326
Illegal EcoRI site found at 4115
Illegal EcoRI site found at 6579
Illegal EcoRI site found at 7518
Illegal XbaI site found at 1813
Illegal XbaI site found at 8965
Illegal XbaI site found at 9775
Illegal XbaI site found at 9889
Illegal SpeI site found at 9930
Illegal PstI site found at 2502
Illegal PstI site found at 3300
Illegal PstI site found at 3795
Illegal PstI site found at 4080
Illegal NgoMIV site found at 2843
Illegal AgeI site found at 425
Illegal AgeI site found at 7131 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 5034
Illegal BsaI site found at 5303
Illegal BsaI.rc site found at 5284
Illegal BsaI.rc site found at 6415
Illegal SapI site found at 4305
Illegal SapI site found at 6865
Illegal SapI.rc site found at 6849
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