LOCUS       sequence-177735-        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      27..615
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     516..535
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     769..786
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     884..901
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     885..901
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1364..1383
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1364..1382
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1383..1407
                     /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             1452..1469
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1470..1490
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      3697..3744
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(3967..3986)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(3992..4011)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4053..4144
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4145..4213,4214..5005)
                     /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(4363..4382)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             5088..6170
                     /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(5107..5126)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
       61 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
      121 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
      181 gtgtagccgt agttagccca ccacttcaag aactctgtag caccgcctac atacctcgct
      241 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
      301 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
      361 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
      421 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
      481 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
      541 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
      601 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
      661 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
      721 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
      781 agcgaggaag cggaaggcga gagtagggaa ctgccaggca tcaaactaag cagaaggccc
      841 ctgacggatg gcctttttgc gtttctacaa actctttctg tgttgtaaaa cgacggccag
      901 tcttaagctc gggccccctg ggcggttctg ataacgagta atcgttaatc cgcaaataac
      961 gtaaaaaccc gcttcggcgg gtttttttat ggggggagtt tagggaaaga gcatttgtca
     1021 gaatatttaa gggcgcctgt cactttgctt gatatatgag aattatttaa ccttataaat
     1081 gagaaaaaag caacgcactt taaataagat acgttgcttt ttcgattgat gaacacctat
     1141 aattaaacta ttcatctatt atttatgatt ttttgtatat acaatatttc tagtttgtta
     1201 aagagaatta agaaaataaa tctcgaaaat aataaaggga aaatcagttt ttgatatcaa
     1261 aattatacat gtcaacgata atacaaaata taatacaaac tataagatgt tatcagtatt
     1321 tattatcatt tagaataaat tttgtgtcgc ccttccgcga aattaatacg actcactata
     1381 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa cttttaggag
     1441 gtaaacatat gcaccatcat catcatcacg agaacttata ctttcaaggc atgagcagcg
     1501 ttacctgggc accgggcaat tacccgagca cccgccgttc cgatcatgtg gacacgtatc
     1561 aaagcgcgag caagggcgaa gtcccggtgc cggatccgta tcagtggctg gaagagagca
     1621 ccgacgaagt cgacaagtgg acgacggcac aagcggacct ggcacagtct tatctggacc
     1681 aaaacgctga cattcaaaaa ctggccgaga aatttcgtgc gagccgtaat tacgccaagt
     1741 tctctgcacc gactctgttg gacgacggtc actggtattg gttctataac cgtggcttgc
     1801 agagccaaag cgtcttgtac agatctaaag aaccagccct gccggacttc tctaagggcg
     1861 acgataacgt tggtgacgtt ttctttgatc cgaatgtgct ggcagcggat ggtagcgcgg
     1921 gcatggtcct gtgcaaattc tcgccggacg gtaaattctt tgcctatgca gttagccacc
     1981 tgggtggcga ttactccacg atttacgtgc gttccacgag cagcccgctg agccaggcga
     2041 gcgtcgcgca aggtgtggac ggtcgtctga gcgacgaagt caaatggttc aagtttagca
     2101 ccattatttg gactaaagat agcaagggct tcctgtacca gcgctatccg gctcgcgagc
     2161 gtcacgaggg tacccgcagc gaccgtaatg ccatgatgtg ttaccacaaa gtcggcacga
     2221 cccaagaaga agatattatt gtgtatcagg ataatgaaca tcctgagtgg atctatggtg
     2281 cggatacctc cgaagatggc aagtacctgt acctgtacca gtttaaagat accagcaaga
     2341 aaaacctgtt gtgggtcgcc gagctggacg aagatggcgt taagagcggt attcactggc
     2401 gtaaagttgt taacgaatac gcagcagact acaatatcat tacgaaccac ggtagcctgg
     2461 tctacattaa gacgaacctg aatgcaccgc aatataaagt tattaccatc gacctgtcca
     2521 aagacgagcc agagatccgt gacttcatcc cggaagaaaa agatgctaaa ctggcgcaag
     2581 ttaattgtgc gaacgaagag tatttcgtcg ctatttacaa gcgtaacgtt aaagatgaaa
     2641 tctacctgta ctcgaaggct ggtgttcagc tgacccgtct ggcgccggat ttcgtgggtg
     2701 cggcaagcat tgcgaatcgt caaaaacaga cccacttttt tctgaccctg agcggtttta
     2761 acaccccggg tacgatcgct cgttatgatt ttaccgcgcc ggaaacccag cgctttagca
     2821 tcctgcgtac cactaaagtc aacgaactgg atcctgatga cttcgagagc acgcaagtgt
     2881 ggtacgagag caaggacggt acgaaaatcc cgatgttcat cgttcgtcac aagtccacca
     2941 agtttgacgg taccgcggct gcgatccaat atggttacgg cggcttcgcc acctccgcgg
     3001 atccattctt cagcccgatc attctgacgt ttctgcagac ctacggtgcg atttttgccg
     3061 taccgagcat ccgtggtggc ggtgagttcg gtgaggaatg gcataagggc ggccgccgcg
     3121 aaaccaaagt gaacacgttt gacgatttca ttgcggccgc ccaatttctg gtgaagaaca
     3181 aatatgcggc tccgggtaaa gttgcaatta atggtgcgag caacggcggc ctgctggtga
     3241 tgggctctat cgtccgtgcg cctgagggta cgttcggtgc ggcagtgcca gagggcggtg
     3301 tggcggatct gcttaagttc cacaaattta cgggtggcca ggcgtggatc agcgagtatg
     3361 gtaatccgtc tatccctgaa gaatttgatt acatctatcc gctgagcccg gttcacaatg
     3421 tccgcaccga caaagttatg ccggcaaccc tgattaccgt gaatatcggt gatggccgtg
     3481 tggttccgat gcacagcttc aagttcatcg cgactttgca gcataacgtt ccgcagaatc
     3541 cgcatccgct gctgattaag attgataaat cgtggttggg tcacggcatg ggtaagccga
     3601 ccgacaaaaa tgtgaaagac gccgccgaca agtggggttt catcgcacgc gccctgggtc
     3661 tggaactgaa aacggtcgag tgatgaaagc ttccccctag cataacccct tggggcctct
     3721 aaacgggtct tgaggggttt tttgcccctg agacgcgtca atcgagttcg tacctaaggg
     3781 cgacaccccc taattagccc gggcgaaagg cccagtcttt cgactgagcc tttcgtttta
     3841 tttgatgcct ggcagttccc tactctcgca tggggagtcc ccacactacc atcggcgcta
     3901 cggcgtttca cttctgagtt cggcatgggg tcaggtggga ccaccgcgct actgccgcca
     3961 ggcaaacaag gggtgttatg agccatattc aggtataaat gggctcgcga taatgttcag
     4021 aattggttaa ttggttgtaa cactgacccc tatttgttta tttttctaaa tacattcaaa
     4081 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     4141 gaatatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     4201 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     4261 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     4321 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     4381 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     4441 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     4501 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     4561 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     4621 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     4681 gatgcctgta gcgatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     4741 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     4801 gcgctcggcc cttccggctg gctggtttat tgctgataaa tccggagccg gtgagcgtgg
     4861 ttctcgcggt atcatcgcag cgctggggcc agatggtaag ccctcccgta tcgtagttat
     4921 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     4981 tgcctcactg attaagcatt ggtaagcggc gcgccatcga atggcgcaaa acctttcgcg
     5041 gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatatg aaaccagtaa
     5101 cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga
     5161 accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc
     5221 tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg
     5281 gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat
     5341 ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg
     5401 aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta
     5461 actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg
     5521 cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgagg
     5581 acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt
     5641 tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc
     5701 tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg
     5761 gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg
     5821 ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg
     5881 gtgcggatat ctcggtagtg ggatacgacg ataccgaaga tagctcatgt tatatcccgc
     5941 cgttaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc
     6001 tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgccagtc tcactggtga
     6061 aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     6121 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga ctcatgacca
     6181 aaatccctta acgtgagtta cgcgcgcgtc gttccactga gcgtcagac
//
