Difference between revisions of "Part:BBa K1159106"

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This parts encodes are fusion protein consisting the photosensitive core of PhyB (2-908 AA) at the N-terminus and the C-terminal half of TEV Protease at its C-terminus. Both parts are seperated by a 36 amino acids long flexible linker. This part is flanked by RFC[25] pre- and suffix to allow further protein fusions.
 
This parts encodes are fusion protein consisting the photosensitive core of PhyB (2-908 AA) at the N-terminus and the C-terminal half of TEV Protease at its C-terminus. Both parts are seperated by a 36 amino acids long flexible linker. This part is flanked by RFC[25] pre- and suffix to allow further protein fusions.
  
By expressing this part together with the second essential protein [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1159107 BBa_K1159107] or [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1159108 BBa_K1159108] that contains the part which interacts with PhyB (PIF3 or PIF6) and the N-terminal half of the splitted TEV Protease, they form together a light-switchable TEV Protease system which can be activated by red light and inactivated by far-red light.
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By expressing this part together with its interaction partner [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1159107 BBa_K1159107] or [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1159108 BBa_K1159108] that contains the part which interacts with PhyB (PIF3 or PIF6) and the N-terminal half of the splitted TEV Protease, they form together a light-switchable TEV Protease system which can be activated by red light and inactivated by far-red light.
  
 
'''''Note: PhyB requires Phycocyanobilin or Phytochromobilin as a cofactor to fulfill its function as photoswitchable protein; you can reach it by adding exogenous Phycocyanobilin/Phytochromobilin in the medium or by inserting genes for the Phycocyanobilin/Phytochromobilin synthesis. See here for more information: [http://2012.igem.org/Team:TU_Munich/Project/Light_Switchable_Promoter#Biosynthesis_of_Phycocyanobilin Click here] '''''  
 
'''''Note: PhyB requires Phycocyanobilin or Phytochromobilin as a cofactor to fulfill its function as photoswitchable protein; you can reach it by adding exogenous Phycocyanobilin/Phytochromobilin in the medium or by inserting genes for the Phycocyanobilin/Phytochromobilin synthesis. See here for more information: [http://2012.igem.org/Team:TU_Munich/Project/Light_Switchable_Promoter#Biosynthesis_of_Phycocyanobilin Click here] '''''  
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K1159106 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K1159106 SequenceAndFeatures</partinfo>
 
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<html><!--- Please copy this table containing parameters for BBa_ at the end of the parametrs section ahead of the references. ---><style type="text/css">table#AutoAnnotator {border:1px solid black; width:100%; border-collapse:collapse;} th#AutoAnnotatorHeader { border:1px solid black; width:100%; background-color: rgb(221, 221, 221);} td.AutoAnnotator1col { width:100%; border:1px solid black; } span.AutoAnnotatorSequence { font-family:'Courier New', Arial; } td.AutoAnnotatorSeqNum { text-align:right; width:2%; } td.AutoAnnotatorSeqSeq { width:98% } td.AutoAnnotatorSeqFeat1 { width:3% } td.AutoAnnotatorSeqFeat2a { width:27% } td.AutoAnnotatorSeqFeat2b { width:97% } td.AutoAnnotatorSeqFeat3 { width:70% } table.AutoAnnotatorNoBorder { border:0px; width:100%; border-collapse:collapse; } table.AutoAnnotatorWithBorder { border:1px solid black; width:100%; border-collapse:collapse; } td.AutoAnnotatorOuterAmino { border:0px solid black; width:20% } td.AutoAnnotatorInnerAmino { border:1px solid black; width:50% } td.AutoAnnotatorAminoCountingOuter { border:1px solid black; width:40%;  } td.AutoAnnotatorBiochemParOuter { border:1px solid black; width:60%; } td.AutoAnnotatorAminoCountingInner1 { width: 7.5% } td.AutoAnnotatorAminoCountingInner2 { width:62.5% } td.AutoAnnotatorAminoCountingInner3 { width:30% } td.AutoAnnotatorBiochemParInner1 { width: 5% } td.AutoAnnotatorBiochemParInner2 { width:55% } td.AutoAnnotatorBiochemParInner3 { width:40% } td.AutoAnnotatorCodonUsage1 { width: 3% } td.AutoAnnotatorCodonUsage2 { width:14.2% } td.AutoAnnotatorCodonUsage3 { width:13.8% } td.AutoAnnotatorAlignment1 { width: 3% } td.AutoAnnotatorAlignment2 { width: 10% } td.AutoAnnotatorAlignment3 { width: 87% } td.AutoAnnotatorLocalizationOuter {border:1px solid black; width:40%} td.AutoAnnotatorGOOuter {border:1px solid black; width:60%} td.AutoAnnotatorLocalization1 { width: 7.5% } td.AutoAnnotatorLocalization2 { width: 22.5% } td.AutoAnnotatorLocalization3 { width: 70% } td.AutoAnnotatorGO1 { width: 5% } td.AutoAnnotatorGO2 { width: 35% } td.AutoAnnotatorGO3 { width: 60% } td.AutoAnnotatorPredFeat1 { width:3% } td.AutoAnnotatorPredFeat2a { width:27% } td.AutoAnnotatorPredFeat3 { width:70% } div.AutoAnnotator_trans { position:absolute; background:rgb(11,140,143); background-color:rgba(11,140,143, 0.8); height:5px; top:100px; } div.AutoAnnotator_sec_helix { position:absolute; background:rgb(102,0,102); background-color:rgba(102,0,102, 0.8); height:5px; top:110px; } div.AutoAnnotator_sec_strand { position:absolute; background:rgb(245,170,26); background-color:rgba(245,170,26, 1); height:5px; top:110px; } div.AutoAnnotator_acc_buried { position:absolute; background:rgb(89,168,15); background-color:rgba(89,168,15, 0.8); height:5px; top:120px; } div.AutoAnnotator_acc_exposed { position:absolute; background:rgb(0, 0, 255); background-color:rgba(0, 0, 255, 0.8); height:5px; top:120px; } div.AutoAnnotator_dis { position:absolute; text-align:center; font-family:Arial,Helvetica,sans-serif; background:rgb(255, 200, 0); background-color:rgba(255, 200, 0, 1); height:16px; width:16px; top:80px; border-radius:50%; } </style><div id='AutoAnnotator_container_1381260103944'><table id="AutoAnnotator"><tr><!-- Time stamp in ms since 1/1/1970 1381260103944 --><th id="AutoAnnotatorHeader" colspan="2">Protein data table for BioBrick <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_<!------------------------Enter BioBrick number here------------------------>">BBa_<!------------------------Enter BioBrick number here------------------------></a> automatically created by the <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">BioBrick-AutoAnnotator</a> version 1.0</th></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Nucleotide sequence</strong> in <strong>RFC 25</strong>, so ATGGCCGGC and ACCGGT were added (in italics) to the 5' and 3' ends: (underlined part encodes the protein)<br><span class="AutoAnnotatorSequence">&nbsp;<u><i>ATGGCCGGC</i>GTTTCCGGA&nbsp;...&nbsp;CTCATGAAT<i>ACCGGT</i></u></span><br>&nbsp;<strong>ORF</strong> from nucleotide position -8 to 3201 (excluding stop-codon)</td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid sequence:</strong> (RFC 25 scars in shown in bold, other sequence features underlined; both given below)<br><span class="AutoAnnotatorSequence"><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqNum">1&nbsp;<br>101&nbsp;<br>201&nbsp;<br>301&nbsp;<br>401&nbsp;<br>501&nbsp;<br>601&nbsp;<br>701&nbsp;<br>801&nbsp;<br>901&nbsp;<br>1001&nbsp;</td><td class="AutoAnnotatorSeqSeq">MAGVSGVGGSGGGRGGGRGGEEEPSSSHTPNNRRGGEQAQSSGTKSLRPRSNTESMSKAIQQYTVDARLHAVFEQSGESGKSFDYSQSLKTTTYGSSVPE<br>QQITAYLSRIQRGGYIQPFGCMIAVDESSFRIIGYSENAREMLGIMPQSVPTLEKPEILAMGTDVRSLFTSSSSILLERAFVAREITLLNPVWIHSKNTG<br>KPFYAILHRIDVGVVIDLEPARTEDPALSIAGAVQSQKLAVRAISQLQALPGGDIKLLCDTVVESVRDLTGYDRVMVYKFHEDEHGEVVAESKRDDLEPY<br>IGLHYPATDIPQASRFLFKQNRVRMIVDCNATPVLVVQDDRLTQSMCLVGSTLRAPHGCHSQYMANMGSIASLAMAVIINGNEDDGSNVASGRSSMRLWG<br>LVVCHHTSSRCIPFPLRYACEFLMQAFGLQLNMELQLALQMSEKRVLRTQTLLCDMLLRDSPAGIVTQSPSIMDLVKCDGAAFLYHGKYYPLGVAPSEVQ<br>IKDVVEWLLANHADSTGLSTDSLGDAGYPGAAALGDAVCGMAVAYITKRDFLFWFRSHTAKEIKWGGAKHHPEDKDDGQRMHPRSSFQAFLEVVKSRSQP<br>WETAEMDAIHSLQLILRDSFKESEAAMNSKVVDGVVQPCRDMAGEQGIDELGAVAREMVRLIETATVPIFAVDAGGCINGWNAKIAELTGLSVEEAMGKS<br>LVSDLIYKENEATVNKLLSRALRGDEEKNVEVKLKTFSPELQGKAVFVVVNACSSKDYLNNIVGVCFVGQDVTSQKIVMDKFINIQGDYKAIVHSPNPLI<br>PPIFAADENTCCLEWNMAMEKLTGWSRSEVIGKMIVGEVFGSCCMLKGPDALTKFMIVLHNAIGGQDTDKFPFPFFDRNGKFVQALLTANKRVSLEGKVI<br>GAFCFLQIPS<b>TG</b>GGSAGGSGSGSSGGSSGASGTGTAGGTGSGSGTGSG<b>TG</b>KSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFIVGIHS<br>ASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPEEPFQPVKEATQLMNTG*</td></tr></table></span></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Sequence features:</strong> (with their position in the amino acid sequence, see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software/FeatureList">list of supported features</a>)<table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorSeqFeat1"></td><td class="AutoAnnotatorSeqFeat2a">RFC25 scar (shown in bold):&nbsp;</td><td class="AutoAnnotatorSeqFeat3">911 to 912, 949 to 950</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Amino acid composition:</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Ala (A)</td><td class="AutoAnnotatorInnerAmino">81 (7.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Arg (R)</td><td class="AutoAnnotatorInnerAmino">48 (4.5%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asn (N)</td><td class="AutoAnnotatorInnerAmino">38 (3.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Asp (D)</td><td class="AutoAnnotatorInnerAmino">55 (5.1%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Cys (C)</td><td class="AutoAnnotatorInnerAmino">22 (2.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gln (Q)</td><td class="AutoAnnotatorInnerAmino">47 (4.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Glu (E)</td><td class="AutoAnnotatorInnerAmino">60 (5.6%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Gly (G)</td><td class="AutoAnnotatorInnerAmino">106 (9.9%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">His (H)</td><td class="AutoAnnotatorInnerAmino">23 (2.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ile (I)</td><td class="AutoAnnotatorInnerAmino">55 (5.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Leu (L)</td><td class="AutoAnnotatorInnerAmino">84 (7.9%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Lys (K)</td><td class="AutoAnnotatorInnerAmino">51 (4.8%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Met (M)</td><td class="AutoAnnotatorInnerAmino">36 (3.4%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Phe (F)</td><td class="AutoAnnotatorInnerAmino">43 (4.0%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Pro (P)</td><td class="AutoAnnotatorInnerAmino">45 (4.2%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Ser (S)</td><td class="AutoAnnotatorInnerAmino">102 (9.5%)</td></tr></table></td><td class="AutoAnnotatorOuterAmino"><table class="AutoAnnotatorWithBorder"><tr><td class="AutoAnnotatorInnerAmino">Thr (T)</td><td class="AutoAnnotatorInnerAmino">56 (5.2%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Trp (W)</td><td class="AutoAnnotatorInnerAmino">14 (1.3%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Tyr (Y)</td><td class="AutoAnnotatorInnerAmino">22 (2.1%)</td></tr><tr><td class="AutoAnnotatorInnerAmino">Val (V)</td><td class="AutoAnnotatorInnerAmino">82 (7.7%)</td></tr></table></td></tr></table></td></tr><tr><td class="AutoAnnotatorAminoCountingOuter"><strong>Amino acid counting</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Total number:</td><td class="AutoAnnotatorAminoCountingInner3">1070</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Positively charged (Arg+Lys):</td><td class="AutoAnnotatorAminoCountingInner3">99 (9.3%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Negatively charged (Asp+Glu):</td><td class="AutoAnnotatorAminoCountingInner3">115 (10.7%)</td></tr><tr><td class="AutoAnnotatorAminoCountingInner1"></td><td class="AutoAnnotatorAminoCountingInner2">Aromatic (Phe+His+Try+Tyr):</td><td class="AutoAnnotatorAminoCountingInner3">102 (9.5%)</td></tr></table></td><td class="AutoAnnotatorBiochemParOuter"><strong>Biochemical parameters</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Atomic composition:</td><td class="AutoAnnotatorBiochemParInner3">C<sub>5078</sub>H<sub>8016</sub>N<sub>1410</sub>O<sub>1566</sub>S<sub>58</sub></td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Molecular mass [Da]:</td><td class="AutoAnnotatorBiochemParInner3">115735.5</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Theoretical pI:</td><td class="AutoAnnotatorBiochemParInner3">5.86</td></tr><tr><td class="AutoAnnotatorBiochemParInner1"></td><td class="AutoAnnotatorBiochemParInner2">Extinction coefficient at 280 nm [M<sup>-1</sup> cm<sup>-1</sup>]:</td><td class="AutoAnnotatorBiochemParInner3">109780 / 111155 (all Cys red/ox)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges</strong>&nbsp;<input type='button' id='hydrophobicity_charge_button' onclick='show_or_hide_plot_1381260103944()' value='Show'><span id="hydrophobicity_charge_explanation"></span><div id="hydrophobicity_charge_container" style='display:none'><div id="hydrophobicity_charge_placeholder0" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder1" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder2" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder3" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder4" style="width:100%;height:150px"></div><div id="hydrophobicity_charge_placeholder5" style="width:100%;height:150px"></div></div></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Codon usage</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Organism:</td><td class="AutoAnnotatorCodonUsage3"><i>E. coli</i></td><td class="AutoAnnotatorCodonUsage3"><i>B. subtilis</i></td><td class="AutoAnnotatorCodonUsage3"><i>S. cerevisiae</i></td><td class="AutoAnnotatorCodonUsage3"><i>A. thaliana</i></td><td class="AutoAnnotatorCodonUsage3"><i>P. patens</i></td><td class="AutoAnnotatorCodonUsage3">Mammals</td></tr><tr><td class="AutoAnnotatorCodonUsage1"></td><td class="AutoAnnotatorCodonUsage2">Codon quality (<a href="http://en.wikipedia.org/wiki/Codon_Adaptation_Index">CAI</a>):</td><td class="AutoAnnotatorCodonUsage3">good (0.73)</td><td class="AutoAnnotatorCodonUsage3">good (0.74)</td><td class="AutoAnnotatorCodonUsage3">good (0.67)</td><td class="AutoAnnotatorCodonUsage3">good (0.75)</td><td class="AutoAnnotatorCodonUsage3">excellent (0.80)</td><td class="AutoAnnotatorCodonUsage3">good (0.66)</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Alignments</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)<table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorAlignment1"></td><td class="AutoAnnotatorAlignment2">SwissProt:</td><td class="AutoAnnotatorAlignment3"><a href='http://www.uniprot.org/uniprot/P14713'>P14713</a> (99% identity on 915 AAs), <a href='http://www.uniprot.org/uniprot/P42497'>P42497</a> (83% identity on 898 AAs)</td></tr><tr><td class="AutoAnnotatorAlignment1"></td><td class="AutoAnnotatorAlignment2">TrEML:</td><td class="AutoAnnotatorAlignment3"><a href='http://www.uniprot.org/uniprot/B0FWJ8'>B0FWJ8</a> (99% identity on 898 AAs), <a href='http://www.uniprot.org/uniprot/Q5G893'>Q5G893</a> (99% identity on 895 AAs), <a href='http://www.uniprot.org/uniprot/Q5G894'>Q5G894</a> (99% identity on 898 AAs), <a href='http://www.uniprot.org/uniprot/Q5G899'>Q5G899</a> (99% identity on 898 AAs), <a href='http://www.uniprot.org/uniprot/Q5G8A1'>Q5G8A1</a> (99% identity on 898 AAs), <a href='http://www.uniprot.org/uniprot/Q5G8A3'>Q5G8A3</a> (99% identity on 913 AAs), <a href='http://www.uniprot.org/uniprot/Q5G8A4'>Q5G8A4</a> (99% identity on 895 AAs), <a href='http://www.uniprot.org/uniprot/Q5G8A5'>Q5G8A5</a> (99% identity on 913 AAs), <a href='http://www.uniprot.org/uniprot/B0FWI4'>B0FWI4</a> (98% identity on 898 AAs), <a href='http://www.uniprot.org/uniprot/B0FWI5'>B0FWI5</a> (98% identity on 898 AAs)</td></tr><tr><td class="AutoAnnotatorAlignment1"></td><td class="AutoAnnotatorAlignment2">PDB:</td><td class="AutoAnnotatorAlignment3">&nbsp;- </td></tr></table></td></tr><tr><th id='AutoAnnotatorHeader' colspan="2"><strong>Predictions</strong> (obtained from <a href='http://predictprotein.org'>PredictProtein.org</a>)</th></tr><tr><td class="AutoAnnotatorLocalizationOuter"><strong>Subcellular Localization</strong> (reliability in brackets)<table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorLocalization1"></td><td class="AutoAnnotatorLocalization2">Archaea:</td><td class="AutoAnnotatorLocalization3">secreted (100%)</td></tr><tr><td class="AutoAnnotatorLocalization1"></td><td class="AutoAnnotatorLocalization2">Bacteria:</td><td class="AutoAnnotatorLocalization3">secreted (52%)</td></tr><tr><td class="AutoAnnotatorLocalization1"></td><td class="AutoAnnotatorLocalization2">Eukarya:</td><td class="AutoAnnotatorLocalization3">cytosol (39%)</td></tr></table></td><td class="AutoAnnotatorGOOuter"><strong>Gene Ontology</strong> (reliability in brackets)<br><table class="AutoAnnotatorNoBorder"><tr><td class='AutoAnnotatorGO1'></td><td class='AutoAnnotatorGO2'>Molecular Function Ontology:</td><td class='AutoAnnotatorGO3'><a href='http://amigo.geneontology.org/cgi-bin/amigo/term_details?term=GO:0009883'>GO:0009883</a> (70%), <a href='http://amigo.geneontology.org/cgi-bin/amigo/term_details?term=GO:0042802'>GO:0042802</a> (26%)</td></tr><tr><td class='AutoAnnotatorGO1'></td><td class='AutoAnnotatorGO2'>Biological Process Ontology:</td><td class='AutoAnnotatorGO3'><a href='http://amigo.geneontology.org/cgi-bin/amigo/term_details?term=GO:0006350'>GO:0006350</a> (30%), <a href='http://amigo.geneontology.org/cgi-bin/amigo/term_details?term=GO:0006355'>GO:0006355</a> (29%)</td></tr><tr><td class='AutoAnnotatorGO1'> </td><td class='AutoAnnotatorGO2'> </td><td class='AutoAnnotatorGO3'>&nbsp;</td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"><strong>Predicted features:</strong><table class="AutoAnnotatorNoBorder"><tr><td class="AutoAnnotatorPredFeat1"></td><td class="AutoAnnotatorPredFeat2a">Disulfid bridges:</td><td class="AutoAnnotatorPredFeat3">&nbsp;- </td></tr><tr><td class="AutoAnnotatorPredFeat1"></td><td class="AutoAnnotatorPredFeat2a">Transmembrane helices:</td><td class="AutoAnnotatorPredFeat3">&nbsp;- </td></tr></table></td></tr><tr><td class="AutoAnnotator1col" colspan="2"> The BioBrick-AutoAnnotator was created by <a href="http://2013.igem.org/Team:TU-Munich">TU-Munich 2013</a> iGEM team. For more information please see the <a href="http://2013.igem.org/Team:TU-Munich/Results/Software">documentation</a>.<br>If you have any questions, comments or suggestions, please leave us a <a href="http://2013.igem.org/Team:TU-Munich/Results/AutoAnnotator">comment</a>.</td></tr></table></div><br><!-- IMPORTANT: DON'T REMOVE THIS LINE, OTHERWISE NOT SUPPORTED FOR IE BEFORE 9 --><!--[if lte IE 8]><script language="javascript" type="text/javascript" src="excanvas.min.js"></script><![endif]--><script type='text/javascript' src='http://code.jquery.com/jquery-1.10.0.min.js'></script><script type='text/javascript' src='http://2013.igem.org/Team:TU-Munich/Flot.js?action=raw&ctype=text/js'></script><script>function show_or_hide_plot_1381260103944(){hydrophobicity_datapoints = 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= [];trans_datapoints = [];sec_helix_datapoints = [[139,143],[147,150],[165,168],[172,182],[230,248],[255,269],[311,320],[358,366],[414,459],[463,465],[470,476],[498,510],[531,537],[586,596],[605,609],[640,664],[684,688],[693,696],[711,723],[774,796],[818,822],[827,831],[851,864],[911,931],[935,939],[943,957],[1016,1024],[1060,1067]];sec_strand_datapoints = [[60,63],[69,73],[121,126],[131,135],[192,196],[202,209],[213,219],[274,281],[287,292],[323,328],[335,336],[370,380],[398,400],[401,405],[481,486],[488,492],[517,519],[540,546],[551,556],[560,565],[614,625],[631,639],[668,672],[677,681],[731,740],[745,756],[762,770],[802,806],[811,815],[871,877],[882,893],[899,907],[971,977],[987,989],[994,999],[1030,1035],[1041,1043],[1046,1049]];acc_exposed_datapoints = 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= 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'<br>&nbsp;<input type=\'checkbox\' id=\'trans_checkbox\' checked=\'checked\'>&nbsp;Predicted transmembrane helices (<img src=\'https://static.igem.org/mediawiki/2013/7/78/TUM13_trans_icon.png\' alt=\'turquois bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'sec_checkbox\' checked=\'checked\'>&nbsp;Predicted secondary structure: Helices (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_helix_icon.png\' alt=\'violet bars\' height=\'10\'></img>) and beta-strands (<img src=\'https://static.igem.org/mediawiki/2013/b/bf/TUM13_strand_icon.png\' alt=\'yellow bars\' height=\'10\'></img>)';description_html = description_html + '<br>&nbsp;<input type=\'checkbox\' id=\'acc_checkbox\' checked=\'checked\'>&nbsp;Predicted solvent accessability: Exposed (<img src=\'https://static.igem.org/mediawiki/2013/1/16/TUM13_exposed_icon.png\' alt=\'blue bars\' height=\'10\'></img>) and buried (<img 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<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Latest revision as of 19:23, 8 October 2013

PhyB(2-908 AA)_36AALinker_C-TEV-Protease (First part for a light-switchable TEV-Protease) in RFC[25]

This parts encodes are fusion protein consisting the photosensitive core of PhyB (2-908 AA) at the N-terminus and the C-terminal half of TEV Protease at its C-terminus. Both parts are seperated by a 36 amino acids long flexible linker. This part is flanked by RFC[25] pre- and suffix to allow further protein fusions.

By expressing this part together with its interaction partner BBa_K1159107 or BBa_K1159108 that contains the part which interacts with PhyB (PIF3 or PIF6) and the N-terminal half of the splitted TEV Protease, they form together a light-switchable TEV Protease system which can be activated by red light and inactivated by far-red light.

Note: PhyB requires Phycocyanobilin or Phytochromobilin as a cofactor to fulfill its function as photoswitchable protein; you can reach it by adding exogenous Phycocyanobilin/Phytochromobilin in the medium or by inserting genes for the Phycocyanobilin/Phytochromobilin synthesis. See here for more information: [http://2012.igem.org/Team:TU_Munich/Project/Light_Switchable_Promoter#Biosynthesis_of_Phycocyanobilin Click here]

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 487
    Illegal BamHI site found at 569
    Illegal XhoI site found at 520
    Illegal XhoI site found at 539
    Illegal XhoI site found at 2674
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2059
    Illegal BsaI site found at 2465
    Illegal SapI site found at 736
    Illegal SapI.rc site found at 3151

Protein data table for BioBrick BBa_ automatically created by the BioBrick-AutoAnnotator version 1.0
Nucleotide sequence in RFC 25, so ATGGCCGGC and ACCGGT were added (in italics) to the 5' and 3' ends: (underlined part encodes the protein)
 ATGGCCGGCGTTTCCGGA ... CTCATGAATACCGGT
 ORF from nucleotide position -8 to 3201 (excluding stop-codon)
Amino acid sequence: (RFC 25 scars in shown in bold, other sequence features underlined; both given below)

101 
201 
301 
401 
501 
601 
701 
801 
901 
1001 
MAGVSGVGGSGGGRGGGRGGEEEPSSSHTPNNRRGGEQAQSSGTKSLRPRSNTESMSKAIQQYTVDARLHAVFEQSGESGKSFDYSQSLKTTTYGSSVPE
QQITAYLSRIQRGGYIQPFGCMIAVDESSFRIIGYSENAREMLGIMPQSVPTLEKPEILAMGTDVRSLFTSSSSILLERAFVAREITLLNPVWIHSKNTG
KPFYAILHRIDVGVVIDLEPARTEDPALSIAGAVQSQKLAVRAISQLQALPGGDIKLLCDTVVESVRDLTGYDRVMVYKFHEDEHGEVVAESKRDDLEPY
IGLHYPATDIPQASRFLFKQNRVRMIVDCNATPVLVVQDDRLTQSMCLVGSTLRAPHGCHSQYMANMGSIASLAMAVIINGNEDDGSNVASGRSSMRLWG
LVVCHHTSSRCIPFPLRYACEFLMQAFGLQLNMELQLALQMSEKRVLRTQTLLCDMLLRDSPAGIVTQSPSIMDLVKCDGAAFLYHGKYYPLGVAPSEVQ
IKDVVEWLLANHADSTGLSTDSLGDAGYPGAAALGDAVCGMAVAYITKRDFLFWFRSHTAKEIKWGGAKHHPEDKDDGQRMHPRSSFQAFLEVVKSRSQP
WETAEMDAIHSLQLILRDSFKESEAAMNSKVVDGVVQPCRDMAGEQGIDELGAVAREMVRLIETATVPIFAVDAGGCINGWNAKIAELTGLSVEEAMGKS
LVSDLIYKENEATVNKLLSRALRGDEEKNVEVKLKTFSPELQGKAVFVVVNACSSKDYLNNIVGVCFVGQDVTSQKIVMDKFINIQGDYKAIVHSPNPLI
PPIFAADENTCCLEWNMAMEKLTGWSRSEVIGKMIVGEVFGSCCMLKGPDALTKFMIVLHNAIGGQDTDKFPFPFFDRNGKFVQALLTANKRVSLEGKVI
GAFCFLQIPSTGGGSAGGSGSGSSGGSSGASGTGTAGGTGSGSGTGSGTGKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFIVGIHS
ASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPEEPFQPVKEATQLMNTG*
Sequence features: (with their position in the amino acid sequence, see the list of supported features)
RFC25 scar (shown in bold): 911 to 912, 949 to 950
Amino acid composition:
Ala (A)81 (7.6%)
Arg (R)48 (4.5%)
Asn (N)38 (3.6%)
Asp (D)55 (5.1%)
Cys (C)22 (2.1%)
Gln (Q)47 (4.4%)
Glu (E)60 (5.6%)
Gly (G)106 (9.9%)
His (H)23 (2.1%)
Ile (I)55 (5.1%)
Leu (L)84 (7.9%)
Lys (K)51 (4.8%)
Met (M)36 (3.4%)
Phe (F)43 (4.0%)
Pro (P)45 (4.2%)
Ser (S)102 (9.5%)
Thr (T)56 (5.2%)
Trp (W)14 (1.3%)
Tyr (Y)22 (2.1%)
Val (V)82 (7.7%)
Amino acid counting
Total number:1070
Positively charged (Arg+Lys):99 (9.3%)
Negatively charged (Asp+Glu):115 (10.7%)
Aromatic (Phe+His+Try+Tyr):102 (9.5%)
Biochemical parameters
Atomic composition:C5078H8016N1410O1566S58
Molecular mass [Da]:115735.5
Theoretical pI:5.86
Extinction coefficient at 280 nm [M-1 cm-1]:109780 / 111155 (all Cys red/ox)
Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges 
Codon usage
Organism:E. coliB. subtilisS. cerevisiaeA. thalianaP. patensMammals
Codon quality (CAI):good (0.73)good (0.74)good (0.67)good (0.75)excellent (0.80)good (0.66)
Alignments (obtained from PredictProtein.org)
SwissProt:P14713 (99% identity on 915 AAs), P42497 (83% identity on 898 AAs)
TrEML:B0FWJ8 (99% identity on 898 AAs), Q5G893 (99% identity on 895 AAs), Q5G894 (99% identity on 898 AAs), Q5G899 (99% identity on 898 AAs), Q5G8A1 (99% identity on 898 AAs), Q5G8A3 (99% identity on 913 AAs), Q5G8A4 (99% identity on 895 AAs), Q5G8A5 (99% identity on 913 AAs), B0FWI4 (98% identity on 898 AAs), B0FWI5 (98% identity on 898 AAs)
PDB: -
Predictions (obtained from PredictProtein.org)
Subcellular Localization (reliability in brackets)
Archaea:secreted (100%)
Bacteria:secreted (52%)
Eukarya:cytosol (39%)
Gene Ontology (reliability in brackets)
Molecular Function Ontology:GO:0009883 (70%), GO:0042802 (26%)
Biological Process Ontology:GO:0006350 (30%), GO:0006355 (29%)
 
Predicted features:
Disulfid bridges: -
Transmembrane helices: -
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