BBa_K1351007
1
BBa_K1351007
CWB domain of B. subtilis minor autolysin LytE
2014-10-04T11:00:00Z
2015-06-01T12:54:58Z
This part was generated by amplification from ''B. subtilis'' W168 gDNA with the primers listed below, followed by digestion with EcoRI and PstI and ligation into pSB1C3. <br> LytE_ENX_SD_Ngo_fwd: gatcGAATTCgcggccgctTCTAGAgtaaggaggaaGCCGGC ATGAAAAAGCAAATCATTACAGCTACGACAGC <br> LytE_207_SNP_Age_rev: gatcCTGCAGcggccgctACTAGTattaACCGGT CGATGAAGATGAAGCAGGTTTTGAGC
<br> A SpeI site was removed by changing T to C at position 399 without altering the protein sequence by fusion PCR with the following mutagenesis primers: <br>
LytE_SpeI399mut_fwd: GTCCTGAAACTGAAAGGTTCAACcAGTTCAAGCAGCTCCAGCTCATCAAAAG <br>
LytE_SpeI399mut_rev: CTTTTGATGAGCTGGAGCTGCTTGAACTgGTTGAACCTTTCAGTTTCAGGAC
Cell wall binding domain and signal peptide (amino acids 1 to 207) of ''Bacillus subtilis'' minor autolysin LytE (probable peptidoglycan endopeptidase, Uniprot [http://www.uniprot.org/uniprot/P54421 P54421]). This domain has been used for displaying beta-lactamase on the cell surface of ''B. subtilis'' (Chen ''et. al.'', 2008). Used in the [http://2014.igem.org/Team:LMU-Munich/Project BaKillus project] to display pathogen-specific peptides to mediate adhesion of ''B. subtilis'' to pathogens.
This part was generated in a modified version of RFC25, where a strong Shine Dalgarno Sequence (SD) is included, and has the following prefix and suffix:
{|
|prefix with EcoRI, NotI, XbaI, SD and NgoMIV:
|<span style="color:blue">GAATTC</span><span style="color:green">GCGGCCGC</span>T<span style="color:red">TCTAGA</span>GT<u>AAGGAGG</u>A<span style="color:orange">GCCGGC</span>
|-
|suffix with AgeI, SpeI, NotI and PstI:
|<span style="color:orange">ACCGGT</span><u>TAA</u>T<span style="color:red">AC<u>TAG</u><u>T</u></span><u>A</u><span style="color:green"><u>G</u>CGGCCGC</span><span style="color:blue">CTGCAG</span>
|}
Sites of restriction enzymes generating compatible overhangs have the same color:
<span style="color:blue">EcoRI</span> and <span style="color:blue">PstI</span> in blue, <span style="color:green">NotI</span> in green, <span style="color:red">XbaI</span> and <span style="color:red">SpeI</span> in red, <span style="color:orange">NgoMIV</span> and <span style="color:orange">AgeI</span> in orange. Shine-Dalgarno sequence and stop codons are underlined.
false
false
_1726_
4206
20048
9
In stock
false
Due to the N-terminal signal peptide, this part allows only C-terminal fusions of proteins to be displayed on the cell surface.
The CWB domain of LytE consists of three N‐terminal LysM cell wall‐binding motifs, ranging from amino acid 28 to 193. To minimize possible effects of a translational fusion to the CWB domain, the part includes the downstream unstructered region until amino acid 207. An additional linker can be inserted to allow more structural flexibilty.
false
Mona Dotzler
annotation2394083
1
signal peptide
range2394083
1
1
75
annotation2394096
1
LysM
range2394096
1
453
581
annotation2394085
1
LysM
range2394085
1
264
392
annotation2394084
1
LysM
range2394084
1
84
212
BBa_K1351007_sequence
1
atgaaaaagcaaatcattacagctacgacagcagttgttttaggatcgacgttatttgcaggagcggcatctgcacaaagcattaaggtgaaaaaaggcgacacgttatgggatctttcaagaaaatacgacacaacgatcagtaaaattaaatcagagaaccaccttcgttcagacattatttatgtgggacaaactttatcgattaacggcaaatctacaagttcaaaaagcagcagttcttcttcctcttcttctacatacaaagtaaagagcggggacagcctttggaaaatttcaaaaaaatacggcatgacaatcaatgaactgaagaagctgaatggcttaaaatcagatttgcttcgtgttggacaagtcctgaaactgaaaggttcaaccagttcaagcagctccagctcatcaaaagtgtcatcgtcttcaacttctacttataaagtgaagagcggagacagcctttctaaaattgcgagcaaatacggcactacggttagcaaattaaaaagcttaaacggcttaaaatcagatgtaatctatgtcaaccaagtattgaaggtgaaaggaacaagcacaagcagctcaaaacctgcttcatcttcatcg
igem2sbol
1
iGEM to SBOL conversion
Conversion of the iGEM parts registry to SBOL2.1
Chris J. Myers
James Alastair McLaughlin
2017-03-06T15:00:00.000Z