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Part:BBa_K1159010

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

Secretory NanoLuc Luciferase (SERK-SigP_nLuc) in RFC[25] N-Part

NanoLuc Luciferase is engineered ATP-independent luciferase from a deep-sea shrimp which luminescense 2 magnitues higher than these from Renilla reniformis or from Photinus pyralis (firefly). Also the molecular weight of NanoLuc luciferase is twice smaller compared to other luciferase (only 19 kDa). This part encodes for a NanoLuc Luciferase which is fused to a N-terminal signal peptide for secretion (SERK signal peptide from Physcomitrella patens). 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
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 116
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

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 ... ATCCTTGCTACCGGT
 ORF from nucleotide position 5 to 622 (excluding stop-codon)
Amino acid sequence: (RFC 25 scars in shown in bold, other sequence features underlined; both given below)

101 
201 
MPGEVAWWRPLFLIALMPIGVLSNAEGDALNTTGVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRIVLSGENGLKIDIHVIIPYEGL
SGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVTPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWRLCE
RILATG*
Sequence features: (with their position in the amino acid sequence, see the list of supported features)
RFC25 scar (shown in bold): 33 to 34
Amino acid composition:
Ala (A)7 (3.4%)
Arg (R)8 (3.9%)
Asn (N)10 (4.9%)
Asp (D)13 (6.3%)
Cys (C)1 (0.5%)
Gln (Q)7 (3.4%)
Glu (E)10 (4.9%)
Gly (G)25 (12.1%)
His (H)4 (1.9%)
Ile (I)20 (9.7%)
Leu (L)21 (10.2%)
Lys (K)7 (3.4%)
Met (M)4 (1.9%)
Phe (F)9 (4.4%)
Pro (P)9 (4.4%)
Ser (S)7 (3.4%)
Thr (T)13 (6.3%)
Trp (W)5 (2.4%)
Tyr (Y)6 (2.9%)
Val (V)20 (9.7%)
Amino acid counting
Total number:206
Positively charged (Arg+Lys):15 (7.3%)
Negatively charged (Asp+Glu):23 (11.2%)
Aromatic (Phe+His+Try+Tyr):24 (11.7%)
Biochemical parameters
Atomic composition:C1039H1617N267O296S5
Molecular mass [Da]:22745.2
Theoretical pI:4.89
Extinction coefficient at 280 nm [M-1 cm-1]:36440 / 36503 (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 (0.66)good (0.69)good (0.62)excellent (0.85)excellent (0.91)excellent (0.81)
Alignments (obtained from PredictProtein.org)
SwissProt:Q9GV45 (81% identity on 194 AAs)
TrEML: -
PDB: -
Predictions (obtained from PredictProtein.org)
Subcellular Localization (reliability in brackets)
Archaea:secreted (100%)
Bacteria:secreted (56%)
Eukarya:cytosol (7%)
Gene Ontology (reliability in brackets)
Molecular Function Ontology: -
Biological Process Ontology: -
 
Predicted features:
Disulfid bridges: -
Transmembrane helices: -
The BioBrick-AutoAnnotator was created by TU-Munich 2013 iGEM team. For more information please see the documentation.
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[edit]
Categories
//chassis/eukaryote/ppatens
Parameters
None