LOCUS       sequence-192472-        6229 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express wild type prolyl oligopeptidase from Galerina marginata.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6229)
  AUTHORS   Czekster CM, Naismith JH
  TITLE     Kinetic Landscape of a Peptide Bond-Forming Prolyl Oligopeptidase.
  JOURNAL   Biochemistry. 2017 Apr 18;56(15):2086-2095. doi: 
            10.1021/acs.biochem.7b00012. Epub 2017 Apr 7.
  PUBMED    28332820
REFERENCE   2  (bases 1 to 6229)
  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"; doi: 
            "10.1021/acs.biochem.7b00012"; journalName: "Biochemistry"; date: 
            "2017-04-18- 18"; volume: "56"; issue: "15"; pages: "2086-2095"
COMMENT     Sequence Label: pJExpress414_GmPOPB
FEATURES             Location/Qualifiers
     source          1..6229
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      561..1149
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1050..1069
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1303..1320
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1418..1435
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1419..1435
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1898..1917
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1898..1916
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1917..1941
                     /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)."
     CDS             1986..2003
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2004..2024
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      4231..4278
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(4501..4520)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4526..4545)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4587..4678
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4679..4747,4748..5539)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4897..4916)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             join(5622..6229,1..475)
                     /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(5641..5660)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 atatctcact cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact ggagtgccat
       61 gtccggtttt caacaaacca tgcaaatgct gaatgagggc atcgttccca ctgcgatgct
      121 ggttgccaac gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg
      181 cgttggtgcg gatatctcgg tagtgggata cgacgatacc gaagatagct catgttatat
      241 cccgccgtta accaccatca aacaggattt tcgcctgctg gggcaaacca gcgtggaccg
      301 cttgctgcaa ctctctcagg gccaggcggt gaagggcaat cagctgttgc cagtctcact
      361 ggtgaaaaga aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
      421 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgactcat
      481 gaccaaaatc ccttaacgtg agttacgcgc gcgtcgttcc actgagcgtc agaccccgta
      541 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
      601 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
      661 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
      721 ccgtagttag cccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
      781 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
      841 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
      901 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
      961 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     1021 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     1081 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     1141 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     1201 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     1261 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     1321 gaagcggaag gcgagagtag ggaactgcca ggcatcaaac taagcagaag gcccctgacg
     1381 gatggccttt ttgcgtttct acaaactctt tctgtgttgt aaaacgacgg ccagtcttaa
     1441 gctcgggccc cctgggcggt tctgataacg agtaatcgtt aatccgcaaa taacgtaaaa
     1501 acccgcttcg gcgggttttt ttatgggggg agtttaggga aagagcattt gtcagaatat
     1561 ttaagggcgc ctgtcacttt gcttgatata tgagaattat ttaaccttat aaatgagaaa
     1621 aaagcaacgc actttaaata agatacgttg ctttttcgat tgatgaacac ctataattaa
     1681 actattcatc tattatttat gattttttgt atatacaata tttctagttt gttaaagaga
     1741 attaagaaaa taaatctcga aaataataaa gggaaaatca gtttttgata tcaaaattat
     1801 acatgtcaac gataatacaa aatataatac aaactataag atgttatcag tatttattat
     1861 catttagaat aaattttgtg tcgcccttcc gcgaaattaa tacgactcac tataggggaa
     1921 ttgtgagcgg ataacaattc ccctctagaa ataattttgt ttaactttta ggaggtaaac
     1981 atatgcacca tcatcatcat cacgagaact tatactttca aggcatgagc agcgttacct
     2041 gggcaccggg caattacccg agcacccgcc gttccgatca tgtggacacg tatcaaagcg
     2101 cgagcaaggg cgaagtcccg gtgccggatc cgtatcagtg gctggaagag agcaccgacg
     2161 aagtcgacaa gtggacgacg gcacaagcgg acctggcaca gtcttatctg gaccaaaacg
     2221 ctgacattca aaaactggcc gagaaatttc gtgcgagccg taattacgcc aagttctctg
     2281 caccgactct gttggacgac ggtcactggt attggttcta taaccgtggc ttgcagagcc
     2341 aaagcgtctt gtacagatct aaagaaccag ccctgccgga cttctctaag ggcgacgata
     2401 acgttggtga cgttttcttt gatccgaatg tgctggcagc ggatggtagc gcgggcatgg
     2461 tcctgtgcaa attctcgccg gacggtaaat tctttgccta tgcagttagc cacctgggtg
     2521 gcgattactc cacgatttac gtgcgttcca cgagcagccc gctgagccag gcgagcgtcg
     2581 cgcaaggtgt ggacggtcgt ctgagcgacg aagtcaaatg gttcaagttt agcaccatta
     2641 tttggactaa agatagcaag ggcttcctgt accagcgcta tccggctcgc gagcgtcacg
     2701 agggtacccg cagcgaccgt aatgccatga tgtgttacca caaagtcggc acgacccaag
     2761 aagaagatat tattgtgtat caggataatg aacatcctga gtggatctat ggtgcggata
     2821 cctccgaaga tggcaagtac ctgtacctgt accagtttaa agataccagc aagaaaaacc
     2881 tgttgtgggt cgccgagctg gacgaagatg gcgttaagag cggtattcac tggcgtaaag
     2941 ttgttaacga atacgcagca gactacaata tcattacgaa ccacggtagc ctggtctaca
     3001 ttaagacgaa cctgaatgca ccgcaatata aagttattac catcgacctg tccaaagacg
     3061 agccagagat ccgtgacttc atcccggaag aaaaagatgc taaactggcg caagttaatt
     3121 gtgcgaacga agagtatttc gtcgctattt acaagcgtaa cgttaaagat gaaatctacc
     3181 tgtactcgaa ggctggtgtt cagctgaccc gtctggcgcc ggatttcgtg ggtgcggcaa
     3241 gcattgcgaa tcgtcaaaaa cagacccact tttttctgac cctgagcggt tttaacaccc
     3301 cgggtacgat cgctcgttat gattttaccg cgccggaaac ccagcgcttt agcatcctgc
     3361 gtaccactaa agtcaacgaa ctggatcctg atgacttcga gagcacgcaa gtgtggtacg
     3421 agagcaagga cggtacgaaa atcccgatgt tcatcgttcg tcacaagtcc accaagtttg
     3481 acggtaccgc ggctgcgatc caatatggtt acggcggctt cgccacctcc gcggatccat
     3541 tcttcagccc gatcattctg acgtttctgc agacctacgg tgcgattttt gccgtaccga
     3601 gcatccgtgg tggcggtgag ttcggtgagg aatggcataa gggcggccgc cgcgaaacca
     3661 aagtgaacac gtttgacgat ttcattgcgg ccgcccaatt tctggtgaag aacaaatatg
     3721 cggctccggg taaagttgca attaatggtg cgagcaacgg cggcctgctg gtgatgggct
     3781 ctatcgtccg tgcgcctgag ggtacgttcg gtgcggcagt gccagagggc ggtgtggcgg
     3841 atctgcttaa gttccacaaa tttacgggtg gccaggcgtg gatcagcgag tatggtaatc
     3901 cgtctatccc tgaagaattt gattacatct atccgctgag cccggttcac aatgtccgca
     3961 ccgacaaagt tatgccggca accctgatta ccgtgaatat cggtgatggc cgtgtggttc
     4021 cgatgcacag cttcaagttc atcgcgactt tgcagcataa cgttccgcag aatccgcatc
     4081 cgctgctgat taagattgat aaatcgtggt tgggtcacgg catgggtaag ccgaccgaca
     4141 aaaatgtgaa agacgccgcc gacaagtggg gtttcatcgc acgcgccctg ggtctggaac
     4201 tgaaaacggt cgagtgatga aagcttcccc ctagcataac cccttggggc ctctaaacgg
     4261 gtcttgaggg gttttttgcc cctgagacgc gtcaatcgag ttcgtaccta agggcgacac
     4321 cccataatta gcccgggcga aaggcccagt ctttcgactg agcctttcgt tttatttgat
     4381 gcctggcagt tccctactct cgcatgggga gtccccacac taccatcggc gctacggcgt
     4441 ttcacttctg agttcggcat ggggtcaggt gggaccaccg cgctactgcc gccaggcaaa
     4501 caaggggtgt tatgagccat attcaggtat aaatgggctc gcgataatgt tcagaattgg
     4561 ttaattggtt gtaacactga cccctatttg tttatttttc taaatacatt caaatatgta
     4621 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagaatat
     4681 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     4741 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     4801 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     4861 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     4921 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     4981 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     5041 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     5101 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     5161 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     5221 tgtagcgatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     5281 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     5341 ggcccttccg gctggctggt ttattgctga taaatccgga gccggtgagc gtggttctcg
     5401 cggtatcatc gcagcgctgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     5461 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     5521 actgattaag cattggtaag cggcgcgcca tcgaatggcg caaaaccttt cgcggtatgg
     5581 catgatagcg cccggaagag agtcaattca gggtggtgaa tatgaaacca gtaacgttat
     5641 acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg
     5701 ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt
     5761 acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg
     5821 ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg
     5881 ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct
     5941 gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc
     6001 cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat
     6061 ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat gaggacggta
     6121 cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg
     6181 gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataa
//
