LOCUS       sequence-369048-        6081 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SH3-588, a designed poly-proline peptide recruiting 
            catechol O-methyltransferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6081)
  AUTHORS   Park R, Ongpipattanakul C, Nair SK, Bowers AA, Kuhlman B
  TITLE     Designer installation of a substrate recruitment domain to tailor 
            enzyme specificity.
  JOURNAL   Nat Chem Biol. 2023 Apr;19(4):460-467. doi: 
            10.1038/s41589-022-01206-0. Epub 2022 Dec 12.
  PUBMED    36509904
REFERENCE   2  (bases 1 to 6081)
  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: 
            "10.1038/s41589-022-01206-0"; journalName: "Nat Chem Biol"; date: 
            "2023-04"; volume: "19"; issue: "4"; pages: "460-467"
COMMENT     Sequence Label: pMCSG28-SH3-588
FEATURES             Location/Qualifiers
     source          1..6081
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(208..225)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(232..252)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     RBS             1095..1115
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1131..1155
                     /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     complement(1155..1174)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1156..1174)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1262..1281)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1487..1564
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1565..2647
                     /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(1584..1603)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3456..3647
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3641..3663)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3749..3888
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3804..3823
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3903..3920)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4074..4662)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4154..4173)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4833..5624,5625..5693))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5456..5475
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5694..5797)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5964..5983
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccgttaatg atgatgatgg tggtgcccgg cggattggaa
      241 gtacaggttc tcccccggtc cctgataaat agctttctct aaaccatcaa ccactttcat
      301 atattccaga taactcgaat aatgtgtgca ttcgaaggat gaagacccac gcacataagc
      361 aaggaagtct ggtgtgccag gtacaatgac attatctgct aacaggacag tacctttgcg
      421 aagcaatccg catttctcca gaagtaaagt gtcgggtaag tagcggtcct tccaatggtc
      481 caaaaatacc atgtctaaag tatcaacgtc gtattttttt ttaagttgcg gaatcaaatc
      541 ttgggaggcc ccgttcagaa tggtgacctt gtcctgcaag ccagcgaaat taagcatttg
      601 ctgtgtgatt gcggcacagt cgggattgat ttccatggtc agaaggcgcg caccaggttg
      661 aagaaggcgg gccatacgaa ccgccgagta tccacaatat gctcccaatt ccaaaactaa
      721 ggaaggagag tactcgcgaa tcaccgcgtc cataatctgc cctttagcgt cacccacatt
      781 catagcccac tcggcatagg tgcaatactg atcgatggcc tccaaaacac tttgtgggtc
      841 tcctggcttg gcgacaagtt ggacaaactc caagatgcgt tgttccttag taccaccact
      901 ccccccagag ccaccaacgg gcgcaacata attgctcgga atatatcctt gctcaccagt
      961 tgtcaggctt acggcaagcc accagtcgcc ctccgaagaa cgcagaatca caaatttctc
     1021 acctttgtga aaagaaaggt catcttctgt acgggcctca taatcgtaaa gagcgacgaa
     1081 taacatggga gagactcctt cttaaagtta aacaaaatta tttctagagg ggaattgtta
     1141 tccgctcaca attcccctat agtgagtcgt attaatttcg cgggatcgag atcgatctcg
     1201 atcctctacg ccggacgcat cgtggccggc atcaccggcg ccacaggtgc ggttgctggc
     1261 gcctatatcg ccgacatcac cgatggggaa gatcgggctc gccacttcgg gctcatgagc
     1321 gcttgtttcg gcgtgggtat ggtggcaggc cccgtggccg ggggactgtt gggcgccatc
     1381 tccttgcatg caccattcct tgcggcggcg gtgctcaacg gcctcaacct actactgggc
     1441 tgcttcctaa tgcaggagtc gcataaggga gagcgtcgag atcccggaca ccatcgaatg
     1501 gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt
     1561 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac
     1621 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga
     1681 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa
     1741 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat
     1801 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt
     1861 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt
     1921 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc
     1981 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat
     2041 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca
     2101 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc
     2161 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga
     2221 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc
     2281 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga
     2341 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag
     2401 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac
     2461 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt
     2521 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc
     2581 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     2641 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccg ggatctcgac
     2701 cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg ggcatgacta
     2761 tcgtcgccgc acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag
     2821 cgctctgggt cattttcggc gaggaccgct ttcgctggag cgcgacgatg atcggcctgt
     2881 cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc cttcgtcact ggtcccgcca
     2941 ccaaacgttt cggcgagaag caggccatta tcgccggcat ggcggcccca cgggtgcgca
     3001 tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc
     3061 agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga
     3121 cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga
     3181 agtcagcgcc ctgcaccatt atgttccgga tctgcatcgc aggatgctgc tggctaccct
     3241 gtggaacacc tacatctgta ttaacgaagc gctggcattg accctgagtg atttttctct
     3301 ggtcccgccg catccatacc gccagttgtt taccctcaca acgttccagt aaccgggcat
     3361 gttcatcatc agtaacccgt atcgtgagca tcctctctcg tttcatcggt atcattaccc
     3421 ccatgaacag aaatccccct tacacggagg catcagtgac caaacaggaa aaaaccgccc
     3481 ttaacatggc ccgctttatc agaagccaga cattaacgct tctggagaaa ctcaacgagc
     3541 tggacgcgga tgaacaggca gacatctgtg aatcgcttca cgaccacgct gatgagcttt
     3601 accgcagctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     3661 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     3721 cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg
     3781 gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccattg
     3841 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct
     3901 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3961 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4021 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4081 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4141 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4201 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4261 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4321 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4381 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4441 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4501 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4561 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4621 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4681 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4741 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4801 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4861 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4921 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4981 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5041 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5101 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg
     5161 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5221 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5281 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5341 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5401 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5461 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5521 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5581 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5641 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5701 ctttttcaaa ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     5761 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     5821 ctaaattgta agcgttaatg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc
     5881 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag
     5941 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg
     6001 gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta
     6061 cagggcgcgt cccattcgcc a
//
