LOCUS       sequence-190665-        7921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express human Insulin Degrading Enzyme (IDE) in E. coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7921)
  AUTHORS   Shen Y, Joachimiak A, Rosner MR, Tang WJ
  TITLE     Structures of human insulin-degrading enzyme reveal a new substrate 
            recognition mechanism.
  JOURNAL   Nature. 2006 Oct 19;443(7113):870-4. Epub 2006 Oct 11.
  PUBMED    17051221
REFERENCE   2  (bases 1 to 7921)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature."; 
            date: "2006-10-19"; volume: "443(7113)"; pages: "870-4. Epub 2006 
            Oct 11"
COMMENT     Sequence Label: pProEx-IDE-wt
FEATURES             Location/Qualifiers
     source          1..7921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    241..257
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     246..268
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             281..298
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3654..3671)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3824..3910
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4002..4029
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4048..4139
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4140..4208,4209..5000)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4358..4377)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5171..5759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5660..5679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5913..5930
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6007..6435)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6117..6138)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6329..6348
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(6473..6492)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6633..6655
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        6733..6810
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             6811..7893
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(6830..6849)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gaattgatct ggtttgacag cttatcatcg actgcacggt gcaccaatgc ttctggcgtc
       61 aggcagccat cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg
      121 tcgctcaagg cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct
      181 ggcaaatatt ctgaaatgag ctgttgacaa ttaatcatcc ggtccgtata atctgtggaa
      241 ttgtgagcgg ataacaattt cacacaggaa acagaccatg catcaccatc accatcacgc
      301 ggccggtatt cccatgaata atccagccat caagagaata ggaaatcaca ttaccaagtc
      361 tcctgaagac aagcgagaat atcgagggct agagctggcc aatggtatca aagtacttct
      421 tatcagtgat cccaccacgg ataagtcatc agcagcactt gatgtgcaca taggttcatt
      481 gtcggatcct ccaaatattg ctggcttaag tcatttttgt gaacatatgc tttttttggg
      541 aacaaagaaa taccctaaag aaaatgaata cagccagttt ctcagtgagc atgcaggaag
      601 ttcaaatgcc tttactagtg gagagcatac caattactat tttgatgttt ctcatgaaca
      661 cctagaaggt gccctagaca ggtttgcaca gttttttctg tgccccttgt tcgatgaaag
      721 ttgcaaagac agagaggtga atgcagttga ttcagaacat gagaagaatg tgatgaatga
      781 tgcctggaga ctctttcaat tggaaaaagc tacagggaat cctaaacacc ccttcagtaa
      841 atttgggaca ggtaacaaat atactctgga gactagacca aaccaagaag gcattgatgt
      901 aagacaagag ctactgaaat tccattctgc ttactattca tccaacttaa tggctgtttg
      961 tgttttaggt cgagaatctt tagatgactt gactaatctg gtggtaaagt tattttctga
     1021 agtagagaac aaaaatgttc cattgccaga atttcctgaa caccctttcc aagaagaaca
     1081 tcttaaacaa ctttacaaaa tagtacccat taaagatatt aggaatctct atgtgacatt
     1141 tcccatacct gaccttcaga aatactacaa atcaaatcct ggtcattatc ttggtcatct
     1201 cattgggcat gaaggtcctg gaagtctgtt atcagaactt aagtcaaagg gctgggttaa
     1261 tactcttgtt ggtgggcaga aggaaggagc ccgaggtttt atgtttttta tcattaatgt
     1321 ggacttgacc gaggaaggat tattacatgt tgaagatata attttgcaca tgtttcaata
     1381 cattcagaag ttacgtgcag aaggacctca agaatgggtt ttccaagagt gcaaggactt
     1441 gaatgctgtt gcttttaggt ttaaagacaa agagaggcca cggggctata catctaagat
     1501 tgcaggaata ttgcattatt atcccctaga agaggtgctc acagcggaat atttactgga
     1561 agaatttaga cctgacttaa tagagatggt tctcgataaa ctcagaccag aaaatgtccg
     1621 ggttgccata gtttctaaat cttttgaagg aaaaactgat cgcacagaag agtggtatgg
     1681 aacccagtac aaacaagaag ctataccgga tgaagtcatc aagaaatggc aaaatgctga
     1741 cctgaatggg aaatttaaac ttcctacaaa gaatgaattt attcctacga attttgagat
     1801 tttaccgtta gaaaaagagg cgacaccata ccctgctctt attaaggata cagctatgag
     1861 caaactttgg ttcaaacaag atgataagtt ttttttgccg aaggcttgtc tcaactttga
     1921 atttttcagc ccatttgctt atgtggaccc cttgcactgt aacatggcct atttgtacct
     1981 tgagctcctc aaagactcac tcaacgagta tgcatatgca gcagagctag caggcttgag
     2041 ctatgatctc caaaatacca tctatgggat gtatctttca gtgaaaggtt acaatgacaa
     2101 gcagccaatt ttactaaaga agattattga gaaaatggct acctttgaga ttgatgaaaa
     2161 aagatttgaa attatcaaag aagcatatat gcgatctctt aacaatttcc gggctgaaca
     2221 gcctcaccag catgccatgt actacctccg cttgctgatg actgaagtgg cctggactaa
     2281 agatgagtta aaagaagctc tggatgatgt aacccttcct cgccttaagg ccttcatacc
     2341 tcagctcctg tcacggctgc acattgaagc ccttctccat ggaaacataa caaagcaggc
     2401 tgcattagga attatgcaga tggttgaaga caccctcatt gaacatgctc ataccaaacc
     2461 tctccttcca agtcagctgg ttcggtatag agaagttcag ctccctgaca gaggatggtt
     2521 tgtttatcag cagagaaatg aagttcacaa taactgtggc atcgagatat actaccaaac
     2581 agacatgcaa agcacctcag agaatatgtt tctggagctc ttctgtcaga ttatctcgga
     2641 accttgcttc aacaccctgc gcaccaagga gcagttgggc tatatcgtct tcagcgggcc
     2701 acgtcgagct aatggcatac agggcttgag attcatcatc cagtcagaaa agccacctca
     2761 ctacctagaa agcagagtgg aagctttctt aattaccatg gaaaagtcca tagaggacat
     2821 gacagaagag gccttccaaa aacacattca ggcattagca attcgtcgac tagacaaacc
     2881 aaagaagcta tctgctgagt gtgctaaata ctggggagaa atcatctccc agcaatataa
     2941 ttttgacaga gataacactg aggttgcata tttaaagaca cttaccaagg aagatatcat
     3001 caaattctac aaggaaatgt tggcagtaga tgctccaagg agacataagg tatccgtcca
     3061 tgttcttgcc agggaaatgg attcttgtcc tgttgttgga gagttcccat gtcaaaatga
     3121 cataaatttg tcacaagcac cagccttgcc acaacctgaa gtgattcaga acatgaccga
     3181 attcaagcgt ggtctgccac tgtttcccct tgtgaaacca catattaact tcatggctgc
     3241 aaaactctga agattcccca tgcatgggaa agtgcaagtg gatgcattcc tgagtcttcc
     3301 agagcctaag aaaatcatct tggccacttt aatagtttct gattcactat tagagaaaca
     3361 aacaaaaaat tgtcaaatgt cattatgtag aaatattata aatccaaaga aaattacaaa
     3421 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     3481 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     3541 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaac ggaattcctc
     3601 gaggcatgcg gtaccaagct tggctgtttt ggcggatgag agaagatttt cagcctgata
     3661 cagattaaat cagaacgcag aagcggtctg ataaaacaga atttgcctgg cggcagtagc
     3721 gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt
     3781 agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc
     3841 tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag
     3901 taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg
     3961 ggcaggacgc ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga
     4021 tggccttttt gcgtttctac aaactctttt gtttattttt ctaaatacat tcaaatatgt
     4081 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     4141 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     4201 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     4261 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     4321 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     4381 gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     4441 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     4501 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     4561 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     4621 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     4681 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     4741 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     4801 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     4861 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     4921 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     4981 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     5041 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     5101 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     5161 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     5221 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     5281 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     5341 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     5401 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     5461 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     5521 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     5581 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     5641 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     5701 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     5761 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     5821 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     5881 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     5941 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataattt
     6001 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     6061 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     6121 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     6181 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     6241 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     6301 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     6361 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     6421 cgctacaggg cgcgtcccat tcgccattca ggctgctatg gtgcactctc agtacaatct
     6481 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     6541 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     6601 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     6661 accgtcatca ccgaaacgcg cgaggcagca gatcaattcg cgcgcgaagg cgaagcggca
     6721 tgcatttacg ttgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc
     6781 ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca
     6841 gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt
     6901 tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac
     6961 cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt
     7021 ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg
     7081 ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg
     7141 gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac
     7201 caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc
     7261 tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc
     7321 gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt
     7381 tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt
     7441 cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg
     7501 caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg
     7561 ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta
     7621 gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa
     7681 caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc
     7741 caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg
     7801 gcacccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     7861 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagcg
     7921 c
//
