Part:BBa_K801060:Design

(+)-Limonene synthase 1 with Strep-tag and yeast consensus sequence.

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 1683
Illegal SapI.rc site found at 107

Protein data table for BioBrick BBa_K801060 automatically created by the BioBrick-AutoAnnotator version 1.0

Nucleotide sequence in RFC 10: (underlined part encodes the protein)
GTACACAATGCGTCGT ... TTCGAAAAATAA
ORF from nucleotide position 8 to 1705 (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

MRRSANYQPSIWDHDFLQSLNSNYTDEAYKRRAEELRGKVKIAIKDVIEPLDQLDLIDNLQRLGLAHRFETEIRNILNNIYNNNKDYNWRKENLYATSLE
FRLLRQHGYPVSQEVFNGFKDDQGGFICDDFKGILSLHEASYYSLEGESIMEEAWQFTSKHLKEVMISKNMEEDVFVAEQAKRALELPLHWKVPMLEARW
FIHIYERREDKNHLLLELAKMEFNTLQAIYQEELKEISGWWKDTGLGEKLSFARNRLVASFLWSMGIAFEPQFAYCRRVLTISIALITVIDDIYDVYGTL
DELEIFTDAVERWDINYALKHLPGYMKMCFLALYNFVNEFAYYVLKQQDFDLLLSIKNAWLGLIQAYLVEAKWYHSKYTPKLEEYLENGLVSITGPLIIT
ISYLSGTNPIIKKELEFLESNPDIVHWSSKIFRLQDDLGTSSDEIQRGDVPKSIQCYMHETGASEEVARQHIKDMMRQMWKKVNAYTADKDSPLTGTTTE
FLLNLVRMSHFMYLHGDGHGVQNQETIDVGFTLLFQPIPLEDKHMAFTASPGTKGTGAWSHPQFEK*

Sequence features: (with their position in the amino acid sequence, see the list of supported features)

	RFC25 scar (shown in bold):	556 to 557
	Strep-tag II:	559 to 566

Amino acid composition:

Ala (A)	33 (5.8%)
Arg (R)	25 (4.4%)
Asn (N)	27 (4.8%)
Asp (D)	34 (6.0%)

Cys (C)	4 (0.7%)
Gln (Q)	24 (4.2%)
Glu (E)	48 (8.5%)
Gly (G)	29 (5.1%)

His (H)	18 (3.2%)
Ile (I)	39 (6.9%)
Leu (L)	64 (11.3%)
Lys (K)	36 (6.4%)

Met (M)	16 (2.8%)
Phe (F)	28 (4.9%)
Pro (P)	17 (3.0%)
Ser (S)	33 (5.8%)

Thr (T)	26 (4.6%)
Trp (W)	14 (2.5%)
Tyr (Y)	27 (4.8%)
Val (V)	24 (4.2%)

Amino acid counting

	Total number:	566
	Positively charged (Arg+Lys):	61 (10.8%)
	Negatively charged (Asp+Glu):	82 (14.5%)
	Aromatic (Phe+His+Try+Tyr):	87 (15.4%)

Biochemical parameters

	Atomic composition:	C₃₀₀₂H₄₅₈₆N₇₇₈O₈₆₈S₂₀
	Molecular mass [Da]:	66105.3
	Theoretical pI:	5.38
	Extinction coefficient at 280 nm [M^-1 cm^-1]:	117230 / 117480 (all Cys red/ox)

Plot for hydrophobicity, charge, predicted secondary structure, solvent accessability, transmembrane helices and disulfid bridges

Codon usage

	Organism:	E. coli	B. subtilis	S. cerevisiae	A. thaliana	P. patens	Mammals
	Codon quality (CAI):	good (0.71)	good (0.75)	good (0.69)	good (0.78)	excellent (0.80)	good (0.68)

Alignments (obtained from PredictProtein.org)

	SwissProt:	Q8L5K3 (99% identity on 554 AAs), Q93X23 (54% identity on 536 AAs)
	TrEML:	A7BG59 (99% identity on 554 AAs), Q8L5K1 (96% identity on 554 AAs)
	PDB:	1n1z (44% identity on 530 AAs), 1n20 (44% identity on 525 AAs)

Predictions (obtained from PredictProtein.org)

Subcellular Localization (reliability in brackets)

	Archaea:	cytosol (100%)
	Bacteria:	cytosol (95%)
	Eukarya:	nucleus (15%)

Gene Ontology (reliability in brackets)

	Molecular Function Ontology:	GO:0050551 (24%), GO:0034768 (24%)
	Biological Process Ontology:	GO:0071485 (19%), GO:0009611 (18%)

Predicted features:

	Disulfid bridges:	-
	Transmembrane helices:	-

The BioBrick-AutoAnnotator was created by TU-Munich 2013 iGEM team. For more information please see the documentation.
If you have any questions, comments or suggestions, please leave us a comment.

Keywords: limonene synthase, limonene

Abbreviations:

Design Notes

Related BioBrick:

Quality control measures:

Biobrick was sequenced

Backbone:

Backbone name: pSB1C3
Resistance: Cp
Copynumber: high

Protein coding:

BioBrick encodes (+)-limonene synthase
UniProt entry: [http://www.uniprot.org/uniprot/Q8L5K3 Q8L5K3]

Enzymatic activity:

E.C. Number: [http://enzyme.expasy.org/EC/4.2.3.20 4.2.3.20]

Cytotoxicity:

see toxicity assay

Safety notes:

Intellectual property:

Corresponding part author/authors:

Source

Source:

BioBrick: BBa_I742111

Forward Primer: TUM12-LIMS1-fw-cons: 5'- GGATCCTCTAGAGTACACAATGCGTCGTTCAGCAAAC - 3'
Reverse Primer: TUM12-LIMS1-rv: 5'- GGATCCACCGGTGCCTTTGGTGCCAGGAGA - 3'

Organism:

Gene sequence derived from Citrus limon

References

Literature references:

[http://www.ncbi.nlm.nih.gov/pubmed/12084056Pubmed: Lücker, J., El Tamer, M. K., Schwab, W., Verstappen, F. W. A., van der Plas, L. H. W., Bouwmeester, H. J., and Verhoeven, H. A., 2002: Monoterpene biosynthesis in lemon (Citrus limon). cDNA isolation and functional analysis of four monoterpene synthases. Eur J Biochem, 269(13):3160–71.]

Database references:

[http://www.ncbi.nlm.nih.gov/nuccore/21435702 GenBank: source of the sequence]
[http://www.uniprot.org/uniprot/Q8L5K3 Uniprot: (R)-limonene synthase 1]