LOCUS       sequence-254340-        6218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N238A mutation in the mature metQ gene from Neisseria 
            meningitides, without the signal sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6218)
  AUTHORS   Nguyen PT, Lai JY, Kaiser JT, Rees DC
  TITLE     Structures of the Neisseria meningitides methionine-binding protein 
            MetQ in substrate-free form and bound to l- and d-methionine 
            isomers.
  JOURNAL   Protein Sci. 2019 Jul 26. doi: 10.1002/pro.3694.
  PUBMED    31348565
REFERENCE   2  (bases 1 to 6218)
  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: "Protein 
            Sci. 2019 Jul 26. doi: 10.1002/pro.3694."
COMMENT     Sequence Label: NmMetQ N238A
FEATURES             Location/Qualifiers
     source          1..6218
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(27..46)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     187..209
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(203..394)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1203..2285)
                     /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     2247..2266
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2286..2363)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2569..2588
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2672..2691
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2672..2690
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2691..2715
                     /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)."
     RBS             2730..2752
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     sig_peptide     2760..2825
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     CDS             2832..2861
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2877..2891
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     primer_bind     complement(3691..3709)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3705..3752
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3789..4244
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3876..3895)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4086..4107
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4270..4374
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4375..4443,4444..5235)
                     /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     complement(4593..4612)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5406..5994
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5895..5914
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6148..6165
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    join(6180..6218,1..101)
                     /label=bom
                     /note="basis of mobility region from pBR322"
ORIGIN
        1 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
       61 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
      121 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
      181 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
      241 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
      301 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
      361 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
      421 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
      481 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
      541 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
      601 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
      661 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
      721 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
      781 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
      841 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
      901 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
      961 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     1021 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     1081 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     1141 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     1201 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     1261 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     1321 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     1381 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     1441 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     1501 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     1561 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     1621 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     1681 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     1741 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     1801 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     1861 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
     1921 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
     1981 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
     2041 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
     2101 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
     2161 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     2221 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     2281 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     2341 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     2401 taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc
     2461 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     2521 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
     2581 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
     2641 gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga
     2701 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatatacata
     2761 tgaaatacct gctgccgacc gctgctgctg gtctgctgct cctcgctgcc cagccggcga
     2821 tggccatggg ccatcatcat catcatcatc atcatcatca cagcagcggc catatcgacg
     2881 acgacgacaa gcatatgaaa gaaatcgtct tcggcacgac cgtcggcgac ttcggcgata
     2941 tggtcaaaga acaaatccaa gccgagctgg agaaaaaggg ctacaccgtc aaactggtcg
     3001 agtttaccga ctatgtacgc ccgaatctgg cattggctga gggcgagttg gacatcaacg
     3061 tcttccaaca caaaccctat cttgacgact tcaaaaaaga acacaatctg gacatcaccg
     3121 aagtcttcca agtgccgacc gcgcctttgg gactgtaccc gggcaagctg aaatcgctgg
     3181 aagaagtcaa agacggcagc accgtatccg cgcccaacga cccgtccaac ttcgcccgcg
     3241 tcttggtgat gctcgacgaa ctgggttgga tcaaactcaa agacggcatc aatccgttga
     3301 ccgcatccaa agcggacatc gccgagaacc tgaaaaacat caaaatcgtc gagcttgaag
     3361 ccgcgcaact gccgcgtagc cgcgccgacg tggattttgc cgtcgtcaac ggcaactacg
     3421 ccataagcag cggcatgaag ctgaccgaag ccctgttcca agaaccgagc tttgcctatg
     3481 tcgcgtggtc tgccgtcaaa accgccgaca aagacagcca atggcttaaa gacgtaaccg
     3541 aggcctataa ctccgacgcg ttcaaagcct acgcgcacaa acgcttcgag ggctacaaat
     3601 cccctgccgc atggaatgaa ggcgcagcca aataactcga ggatccggct gctaacaaag
     3661 cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg
     3721 gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg
     3781 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     3841 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     3901 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     3961 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     4021 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     4081 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     4141 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     4201 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg
     4261 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     4321 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     4381 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     4441 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     4501 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     4561 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     4621 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     4681 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     4741 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4801 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4861 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
     4921 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4981 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     5041 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     5101 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     5161 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     5221 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     5281 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     5341 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     5401 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     5461 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     5521 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     5581 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     5641 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     5701 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     5761 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5821 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5881 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5941 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     6001 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     6061 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     6121 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     6181 ctgatgcggt attttctcct tacgcatctg tgcggtat
//
