LOCUS       Exported               10763 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pBBR1-aceEF-lpd, complete sequence.
ACCESSION   MH651726
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10763)
  AUTHORS   Yang D, Kim WJ, Yoo SM, Choi JH, Ha SH, Lee MH, Lee SY.
  TITLE     Repurposing type III polyketide synthase as a malonyl-CoA biosensor 
            for metabolic engineering in bacteria
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. (2018) In press
  PUBMED    30232266
REFERENCE   2  (bases 1 to 10763)
  AUTHORS   Yang D, Kim WJ, Yoo SM, Choi JH, Ha SH, Lee MH, Lee SY.
  TITLE     Novel Enzyme-coupled Malonyl-CoA Biosensor based on Type III 
            Polyketide Synthase and Use of the same (KR 10-2018-0066323)
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 10763)
  AUTHORS   Yang D, Kim WJ, Yoo SM, Choi JH, Ha SH, Lee MH, Lee SY.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-JUL-2018) Dept. Chemical and Biomolecular Engineering,
            Korea Advanced Institute of Science and Technology, Daehak-ro 291, 
            Daejeon 34141, South Korea
REFERENCE   4  (bases 1 to 10763)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10763
                     /organism="Cloning vector pBBR1-aceEF-lpd"
                     /mol_type="other DNA"
                     /db_xref="taxon:2364884"
     regulatory      2..30
                     /regulatory_class="promoter"
                     /note="Tac promoter"
     promoter        2..30
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    38..54
                     /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             77..2740
                     /codon_start=1
                     /gene="aceE"
                     /product="pyruvate dehydrogenase E1 component"
                     /label=aceE
                     /protein_id="AYD60213.1"
                     /translation="MSERFPNDVDPIETRDWLQAIESVIREEGVERAQYLIDQLLAEAR
                     KGGVNVAAGTGISNYINTIPVEEQPEYPGNLELERRIRSAIRWNAIMTVLRASKKDLEL
                     GGHMASFQSSATIYDVCFNHFFRARNEQDGGDLVYFQGHISPGVYARAFLEGRLTQEQL
                     DNFRQEVHGNGLSSYPHPKLMPEFWQFPTVSMGLGPIGAIYQAKFLKYLEHRGLKDTSK
                     QTVYAFLGDGEMDEPESKGAITIATREKLDNLVFVINCNLQRLDGPVTGNGKIINELEG
                     IFEGAGWNVIKVMWGSRWDELLRKDTSGKLIQLMNETVDGDYQTFKSKDGAYVREHFFG
                     KYPETAALVADWTDEQIWALNRGGHDPKKIYAAFKKAQETKGKATVILAHTIKGYGMGD
                     AAEGKNIAHQVKKMNMDGVRHIRDRFNVPVSDTDIEKLPYITFPEGSEEHTYLHAQRQK
                     LHGYLPSRQPNFTEKLELPSLQDFGALLEEQSKEISTTIAFVRALNVMLKNKSIKDRLV
                     PIIADEARTFGMEGLFRQIGIYSPNGQQYTPQDREQVAYYKEDEKGQILQEGINELGAG
                     CSWLAAATSYSTNNLPMIPFYIYYSMFGFQRIGDLCWAAGDQQARGFLIGGTSGRTTLN
                     GEGLQHEDGHSHIQSLTIPNCISYDPAYAYEVAVIMHDGLERMYGEKQENVYYYITTLN
                     ENYHMPAMPEGAEEGIRKGIYKLETIEGSKGKVQLLGSGSILRHVREAAEILAKDYGVG
                     SDVYSVTSFTELARDGQDCERWNMLHPLETPRVPYIAQVMNDAPAVASTDYMKLFAEQV
                     RTYVPADDYRVLGTDGFGRSDSRENLRHHFEVDASYVVVAALGELAKRGEIDKKVVADA
                     IAKFNIDADKVNPRLA"
     gene            77..2740
                     /gene="aceE"
                     /label=aceE
     CDS             2755..4647
                     /codon_start=1
                     /gene="aceF"
                     /product="pyruvate dehydrogenase E2 component"
                     /label=aceF
                     /protein_id="AYD60214.1"
                     /translation="MAIEIKVPDIGADEVEITEILVKVGDKVEAEQSLITVEGDKASME
                     VPSPQAGIVKEIKVSVGDKTQTGALIMIFDSADGAADAAPAQAEEKKEAAPAAAPAAAA
                     AKDVNVPDIGSDEVEVTEILVKVGDKVEAEQSLITVEGDKASMEVPAPFAGTVKEIKVN
                     VGDKVSTGSLIMVFEVAGEAGAAAPAAKQEAAPAAAPAPAAGVKEVNVPDIGGDEVEVT
                     EVMVKVGDKVAAEQSLITVEGDKASMEVPAPFAGVVKELKVNVGDKVKTGSLIMIFEVE
                     GAAPAAAPAKQEAAAPAPAAKAEAPAAAPAAKAEGKSEFAENDAYVHATPLIRRLAREF
                     GVNLAKVKGTGRKGRILREDVQAYVKEAIKRAEAAPAATGGGIPGMLPWPKVDFSKFGE
                     IEEVELGRIQKISGANLSRNWVMIPHVTHFDKTDITELEAFRKQQNEEAAKRKLDVKIT
                     PVVFIMKAVAAALEQMPRFNSSLSEDGQRLTLKKYINIGVAVDTPNGLVVPVFKDVNKK
                     GIIELSRELMTISKKARDGKLTAGEMQGGCFTISSIGGLGTTHFAPIVNAPEVAILGVS
                     KSAMEPVWNGKEFVPRLMLPISLSFDHRVIDGADGARFITIINNTLSDIRRLVM"
     gene            2755..4647
                     /gene="aceF"
                     /label=aceF
     regulatory      4671..4699
                     /regulatory_class="promoter"
                     /note="Tac promoter"
     promoter        4671..4699
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    4707..4723
                     /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             4746..6170
                     /codon_start=1
                     /gene="lpd"
                     /product="pyruvate dehydrogenase E3 component"
                     /label=lpd
                     /protein_id="AYD60215.1"
                     /translation="MSTEIKTQVVVLGAGPAGYSAAFRCADLGLETVIVERYNTLGGVC
                     LNVGCIPSKALLHVAKVIEEAKALAEHGIVFGEPKTDIDKIRTWKEKVINQLTGGLAGM
                     AKGRKVKVVNGLGKFTGANTLEVEGENGKTVINFDNAIIAAGSRPIQLPFIPHEDPRIW
                     DSTDALELKEVPERLLVMGGGIIGLEMGTVYHALGSQIDVVEMFDQVIPAADKDIVKVF
                     TKRISKKFNLMLETKVTAVEAKEDGIYVTMEGKKAPAEPQRYDAVLVAIGRVPNGKNLD
                     AGKAGVEVDDRGFIRVDKQLRTNVPHIFAIGDIVGQPMLAHKGVHEGHVAAEVIAGKKH
                     YFDPKVIPSIAYTEPEVAWVGLTEKEAKEKGISYETATFPWAASGRAIASDCADGMTKL
                     IFDKESHRVIGGAIVGTNGGELLGEIGLAIEMGCDAEDIALTIHAHPTLHESVGLAAEV
                     FEGSITDLPNPKAKKK"
     gene            4746..6170
                     /gene="lpd"
                     /label=lpd
     regulatory      6214..6624
                     /regulatory_class="terminator"
                     /note="rrnBT1T2"
     terminator      6419..6505
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6597..6624
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     protein_bind    6645..6666
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(6723..7412)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol resistance protein"
                     /label=cat
                     /protein_id="AYD60216.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQFLCMRPIRKPPL
                     PARWPIH"
     gene            complement(6723..7412)
                     /gene="cat"
                     /label=cat
     promoter        complement(7413..7515)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(7665..8684)
                     /codon_start=1
                     /gene="mob"
                     /product="Mob"
                     /function="required for plasmid mobilization"
                     /label=mob
                     /protein_id="AYD60217.1"
                     /translation="MAAYAIMRCKKLAKMGNVAASLKHAYRERETPNADASRTPENEHW
                     AASSTDEAMGRLRELLPEKRRKDAVLAVEYVMTASPEWWKSASQEQQAAFFEKAHKWLA
                     DKYGADRIVTASIHRDETSPHMTAFVVPLTQDGRLSAKEFIGNKAQMTRDQTTFAAAVA
                     DLGLQRGIEGSKARHTRIQAFYEALERPPVGHVTISPQAVEPRAYAPQGLAEKLGISKR
                     VETPEAVADRLTKAVRQGYEPALQAAAGAREMRKKADQAQETARDLRERLKPVLDALGP
                     LNRDMQAKAAAIIKAVGEKLLTEQREVQRQKQAQRQQERGRAHFPEKCHLTSKKPLLS"
     gene            complement(7665..8684)
                     /gene="mob"
                     /label=mob
     rep_origin      8908..9679
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             9680..10342
                     /codon_start=1
                     /gene="rep"
                     /product="Rep"
                     /function="required for plasmid replication"
                     /label=rep
                     /protein_id="AYD60218.1"
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     gene            9680..10342
                     /gene="rep"
                     /label=rep