LOCUS       sequence-81868-e        1284 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses SpyTag-SpyCatcher mediated cyclized version of TEM-1 
            beta-lactamase (BLA). Cyclization of BLA led to an increase in 
            thermal resilience of >60 degrees celsius..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1284)
  AUTHORS   Schoene C, Fierer JO, Bennett SP, Howarth M
  TITLE     SpyTag/SpyCatcher cyclization confers resilience to boiling on a 
            mesophilic enzyme.
  JOURNAL   Angew Chem Int Ed Engl. 2014 Jun 10;53(24):6101-4. doi: 
            10.1002/anie.201402519. Epub 2014 May 9.
  PUBMED    24817566
REFERENCE   2  (bases 1 to 1284)
  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: 
            "10.1002/anie.201402519"; journalName: "Angew Chem Int Ed Engl"; 
            date: "2014-06-10- 10"; volume: "53"; issue: "24"; pages: "6101-4"
COMMENT     Sequence Label: SpyTag-beta-Lactamase-SpyCatcher
FEATURES             Location/Qualifiers
     source          1..1284
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             13..30
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             40..57
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             67..105
                     /codon_start=1
                     /product="peptide derived from the CnaB2 adhesin domain of 
                     Streptococcus pyogenes FbaB"
                     /label=SpyTag
                     /note="forms a covalent isopeptide bond with the SpyCatcher
                     protein (Zakeri et al., 2012)"
                     /translation="AHIVMVDAYKPTK"
     CDS             127..912
                     /codon_start=1
                     /gene="bla(M)"
                     /product="beta-lactamase lacking the signal sequence"
                     /label=bla(M)
                     /note="allows cytosolic expression of beta-lactamase"
                     /translation="PETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMST
                     FKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMS
                     DNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPAAMATT
                     LRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAA
                     LGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"
     primer_bind     complement(273..292)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             940..1278
                     /codon_start=1
                     /product="truncated version of the CnaB2 adhesin domain of 
                     Streptococcus pyogenes FbaB"
                     /label=SpyCatcher
                     /note="forms a covalent isopeptide bond with the SpyTag 
                     peptide (Zakeri et al., 2012)"
                     /translation="VDTLSGLSSEQGQSGDMTIEEDSATHIKFSKRDEDGKELAGATME
                     LRDSSGKTISTWISDGQVKDFYLYPGKYTFVETAAPDGYEVATAITFTVNEQGQVTVNG
                     KATKGDAHI"
ORIGIN
        1 atgggcagca gccatcatca tcatcatcac agcagcggcc tggtgccgcg cggcagccat
       61 atgggagccc acatcgtgat ggtggacgcc tacaagccga cgaagggttc agggggttcc
      121 ggtcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      181 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      241 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
      301 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
      361 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
      421 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
      481 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
      541 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
      601 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
      661 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
      721 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
      781 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
      841 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
      901 attaagcatt gggggagtgg tggcagcgga ggcgccatgg ttgatacctt atcaggttta
      961 tcaagtgagc aaggtcagtc cggtgatatg acaattgaag aagatagtgc tacccatatt
     1021 aaattctcaa aacgtgatga ggacggcaaa gagttagctg gtgcaactat ggagttgcgt
     1081 gattcatctg gtaaaactat tagtacatgg atttcagatg gacaagtgaa agatttctac
     1141 ctgtatccag gaaaatatac atttgtcgaa accgcagcac cagacggtta tgaggtagca
     1201 actgctatta cctttacagt taatgagcaa ggtcaggtta ctgtaaatgg caaagcaact
     1261 aaaggtgacg ctcatattta atga
//
