Coding

Part:BBa_K1159018

Designed by: Rosario CICCONE   Group: iGEM13_TU-Munich   (2013-05-17)

Membrane-anchored Catechol-2,3-dioxygenase (XylE) in RFC[25] N-Part

This part codes for a membrane-anchored version of Catechol-2,3-dioxygenase (XylE) for the chassis Physcomitrella patens. Please see subparts for for further description. This construct is flanked by RFC[25] N-part prefix and suffix. Note: This means only protein fusions to the C-terminus of this part is possible, adding promoters (typically RFC[10]) into the prefix or terminators/IRES (typically RFC[10]) into the suffix of this part is nevertheless possible.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1038
    Illegal BsaI.rc site found at 1956

Protein data table for BioBrick BBa_ automatically created by the BioBrick-AutoAnnotator version 1.0
Nucleotide sequence in RFC 25 N-Part using the stop codon in the suffix, so ACCGGT was added (in italics) to the 3' end: (underlined part encodes the protein)
 AACAATGCCTGGT ... CTCTACAAAACCGGT
 ORF from nucleotide position 5 to 2032 (excluding stop-codon)
Amino acid sequence: (RFC 25 scars in shown in bold, other sequence features underlined; both given below)
1 
101 
201 
301 
401 
501 
601 		MPGEVAWWRPLFLIALMPIGVLSNAEGDALNTTGNKGVMRPGHVQLRVLDMSKALEHYVELLGLIEMDRDDQGRVYLKAWTEVDKFSLVLREADEPGMDF
MGFKVVDEDALRQLERDLMAYGCAVEQLPAGELNSCGRRVRFQAPSGHHFELYADKEYTGKWGLNDVNPEAWPRDLKGMAAVRFDHALMYGDELPATYDL
FTKVLGFYLAEQVLDENGTRVAQFLSLSTKAHDVAFIHHPEKGRLHHVSFHLETWEDLLRAADLISMTDTSIDIGPTRHGLTHGKTIYFFDPSGNRNEVF
CGGDYNYPDHKPVTWTTDQLGKAIFYHDRILNERFMTVLTTGSAWSHPQFEKGENLYFQSGTGPGQPPFPPPPPFTPPPPQTPNGASGENSTGAIAGGVA
AGAALLFAAPAIGFAWWRRRRPIEATGGGSGGGSGTGSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTL
TYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
FKIRHNIEDGSVQLADHYQQNTPIGDGPVLSPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITHGMDELYKTG*
Sequence features: (with their position in the amino acid sequence, see the list of supported features)
RFC25 scar (shown in bold): 33 to 34, 341 to 342, 362 to 363, 426 to 427, 436 to 437
Strep-tag II: 345 to 352
TEV cleavage site: 354 to 360
Amino acid composition:
Ala (A)46 (6.8%)
Arg (R)30 (4.4%)
Asn (N)27 (4.0%)
Asp (D)46 (6.8%)
Cys (C)5 (0.7%)
Gln (Q)20 (3.0%)
Glu (E)43 (6.4%)
Gly (G)71 (10.5%)
His (H)26 (3.8%)
Ile (I)25 (3.7%)
Leu (L)61 (9.0%)
Lys (K)35 (5.2%)
Met (M)17 (2.5%)
Phe (F)35 (5.2%)
Pro (P)41 (6.1%)
Ser (S)29 (4.3%)
Thr (T)43 (6.4%)
Trp (W)11 (1.6%)
Tyr (Y)24 (3.6%)
Val (V)41 (6.1%)
Amino acid counting
Total number:676
Positively charged (Arg+Lys):65 (9.6%)
Negatively charged (Asp+Glu):89 (13.2%)
Aromatic (Phe+His+Try+Tyr):96 (14.2%)
Biochemical parameters
Atomic composition:C3370H5127N911O998S22
Molecular mass [Da]:75077.6
Theoretical pI:5.53
Extinction coefficient at 280 nm [M-1 cm-1]:96260 / 96573 (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 (0.73)good (0.65)good (0.74)excellent (0.82)good (0.74)
Alignments (obtained from PredictProtein.org)
SwissProt:P06622 (100% identity on 306 AAs), P42212 (98% identity on 237 AAs)
TrEML:D2XKL2 (100% identity on 306 AAs), Q38Q96 (100% identity on 293 AAs), Q7AXV1 (100% identity on 306 AAs), Q7BH44 (100% identity on 306 AAs), Q9L780 (99% identity on 306 AAs), B6UPG7 (98% identity on 237 AAs), Q71RY9 (98% identity on 237 AAs), Q8GHE2 (98% identity on 237 AAs)
PDB:1mpy (100% identity on 306 AAs), 2y0g (98% identity on 226 AAs)
Predictions (obtained from PredictProtein.org)
Subcellular Localization (reliability in brackets)
Archaea:cytosol (100%)
Bacteria:cytosol (51%)
Eukarya:cytosol (40%)
Gene Ontology (reliability in brackets)
Molecular Function Ontology: -
Biological Process Ontology:GO:0019439 (42%), GO:0055114 (29%)
 
Predicted features:
Disulfid bridges: -
Transmembrane helices:398 to 415 going inwards
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Categories
Parameters
None