Difference between revisions of "Part:BBa K801060:Design"

Revision as of 22:11, 7 September 2013

(+)-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: (RFC25 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)

Hydrophobicity and charge plot

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)

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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]

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flot_plot_options = []; flot_plot_options[0] = {		grid: {		borderWidth: {			top: 0,			right: 0,			bottom: 0,			left: 0		}	},	legend: {show: false},	xaxes: [{		min: 0,		max: 200,		ticks: [[0.5, '1'], [24.5, '25'], [49.5, '50'], [74.5, '75'], [99.5, '100'], [124.5, '125'], [149.5, '150'], [174.5, '175'], [199.5, '200']],		tickLength: -5		}],	yaxes: [{		ticks: [[0, '0'], [4.5,'hydro-<br>phobic&nbsp;&nbsp;'], [-4.5,'hydro-<br>philic&nbsp;&nbsp;']],		min: -4.5,		max: +4.5,		font: {		    size: 12,		    lineHeight: 14,		    style: 'italic',		    weight: 'bold',		    family: 'sans-serif',		    variant: 'small-caps',		    color: 'rgba(100,149,237,1)'		}	},	{		ticks: [[0, ''], [1,'positive<br>&nbsp;charge'], [-1,'negative<br>&nbsp;charge']],		position: 'right',		min: -1,		max: 1,		font: {		    size: 12,		    lineHeight: 14,		    style: 'italic',		    weight: 'bold',		    family: 'sans-serif',		    variant: 'small-caps',		    color: 'rgba(255,99,71,1)'		}	}]};var number_of_plots = Math.ceil(amino_sequence.length/200);for ( plot_num = 1 ; plot_num < number_of_plots ; plot_num ++){flot_plot_options[plot_num] = $.extend(true, {} ,flot_plot_options[0]);flot_plot_options[plot_num].xaxes = [{	min: plot_num*200,	max: (plot_num + 1)*200,	ticks: [ [plot_num*200 +   0.5, (plot_num*200 +   1).toString()], 	         [plot_num*200 +  24.5, (plot_num*200 +  25).toString()], 	         [plot_num*200 +  49.5, (plot_num*200 +  50).toString()], 	         [plot_num*200 +  74.5, (plot_num*200 +  75).toString()], 	         [plot_num*200 +  99.5, (plot_num*200 + 100).toString()], 	         [plot_num*200 + 124.5, (plot_num*200 + 125).toString()], 	         [plot_num*200 + 149.5, (plot_num*200 + 150).toString()], 	         [plot_num*200 + 174.5, (plot_num*200 + 175).toString()], 	         [plot_num*200 + 199.5, (plot_num*200 + 200).toString()] ],	tickLength: -5	}];};
+var hydrophobicity_datapoints = 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+</script>
+var flot_plot_options = []; flot_plot_options[0] = {		grid: {		borderWidth: {			top: 0,			right: 0,			bottom: 0,			left: 0		}	},	legend: {show: false},	xaxes: [{		min: 0,		max: 200,		ticks: [[0.5, '1'], [24.5, '25'], [49.5, '50'], [74.5, '75'], [99.5, '100'], [124.5, '125'], [149.5, '150'], [174.5, '175'], [199.5, '200']],		tickLength: -5		}],	yaxes: [{		ticks: [[0, '0'], [4.5,'hydro-<br>phobic&nbsp;&nbsp;'], [-4.5,'hydro-<br>philic&nbsp;&nbsp;']],		min: -4.5,		max: +4.5,		font: {		    size: 12,		    lineHeight: 14,		    style: 'italic',		    weight: 'bold',		    family: 'sans-serif',		    variant: 'small-caps',		    color: 'rgba(100,149,237,1)'		}	},	{		ticks: [[0, ''], [1,'positive<br>&nbsp;charge'], [-1,'negative<br>&nbsp;charge']],		position: 'right',		min: -1,		max: 1,		font: {		    size: 12,		    lineHeight: 14,		    style: 'italic',		    weight: 'bold',		    family: 'sans-serif',		    variant: 'small-caps',		    color: 'rgba(255,99,71,1)'		}	}]};var number_of_plots = Math.ceil(amino_sequence.length/200);for ( plot_num = 1 ; plot_num < number_of_plots ; plot_num ++){flot_plot_options[plot_num] = $.extend(true, {} ,flot_plot_options[0]);flot_plot_options[plot_num].xaxes = [{	min: plot_num*200,	max: (plot_num + 1)*200,	ticks: [ [plot_num*200 +   0.5, (plot_num*200 +   1).toString()], 	         [plot_num*200 +  24.5, (plot_num*200 +  25).toString()], 	         [plot_num*200 +  49.5, (plot_num*200 +  50).toString()], 	         [plot_num*200 +  74.5, (plot_num*200 +  75).toString()], 	         [plot_num*200 +  99.5, (plot_num*200 + 100).toString()], 	         [plot_num*200 + 124.5, (plot_num*200 + 125).toString()], 	         [plot_num*200 + 149.5, (plot_num*200 + 150).toString()], 	         [plot_num*200 + 174.5, (plot_num*200 + 175).toString()], 	         [plot_num*200 + 199.5, (plot_num*200 + 200).toString()] ],	tickLength: -5	}];};
 alert("Here");
 function show_or_hide_plot(){	try {		if( $('#hydrophobicity_charge_button').val() =='Show' ){			$('#hydrophobicity_charge_container').css('display','block');			for (plot_num = 0 ; plot_num < number_of_plots ; plot_num ++){				$.plot('#hydrophobicity_charge_placeholder'+ plot_num.toString(),						[{ 							color: 'rgba(100,149,237,1)',							data: hydrophobicity_datapoints,							label: 'Hydrophobicity',							lines: { show: true, fill: true, fillColor: 'rgba(100,149,237,0.1)' },							yaxis: 1						}, {							color: 'rgba(255,99,71,1)',							data: charge_datapoints,							label: 'Charge',							lines: { show: true, fill: true, fillColor: 'rgba(255,99,71,0.1)' },							yaxis: 2						}],						flot_plot_options[plot_num] );			}			$('#hydrophobicity_charge_button').val('Hide');			$('#hydrophobicity_charge_explanation').text(': Moving average over 5 amino acids for hydrophobicity (blue) and charge (red)');		}		else{			$('#hydrophobicity_charge_container').css('display','none');			$('#hydrophobicity_charge_button').val('Show');			$('#hydrophobicity_charge_explanation').text('');		}	}	catch(err){		txt='There was an error with the button controlling the visibility of the plot.