desABCD cluster producing biosynthesis pathway to make Desferrioxamine B
We used 7 previous biobricks to design this one : - BBa_B0034 - BBa_I0500 - BBa_K1951000 - BBa_K1951001 - BBa_K1951002 - BBa_K1951003
Desferrioxamine B is a siderophore produced in Streptomyces coelicolor to catch vital irons on the extracellular medium. In consequence, this molecule which have a greatly high affinity for iron is secreted in the external medium and bring back inside the bacteria full of iron mol??cules. Previous work showed that siderophore which are molecule secreted by bacteria to catch iron are able to catch other metals by default. Especifically, many articles showed a high affinity of Desferrioxamine B ( produce by ''Streptomyces coelicolor'') for tetravalent metal ions and more specifically platinum. These results encouraged us to make a biobrick coding the sequence corresponding to the 4 enzymes involved on the metabolic pathway of Desferrioxamine B. ''E. coli'' was used to produce this biobrick. Produce a gram positive bacteria pathway in a gram negative bacteria is restrictive, considering the risk of toxicity for the conductress bacteria. To counter this potential issue, we regulate trasnscription using the control of an inductible promotor (pBAD/araC). Our strategy for the siderophore mobilisation results in two main steps : 1. Transformation of E.Coli with an inductible plasmide holding the ''des'' cluster 2. Induction and production of the desferrioxamine B
There is no any design consideration for this biobrick.
RBS (Elowitz 1999) -- defines RBS efficiency
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix. No secondary structures are formed in the given RBS region. Users should check for secondary structures induced in the RBS by upstream and downstream elements in the +50 to -50 region, as such structures will greatly affect the strength of the RBS. Contact info for this part: Brian Chow
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
desA, lysine decarboxylase coding sequence
This part comes from the genome sequencing of Streptomyces coelicolor. The following website enabled to find the gene reference and its chromosomal position. http://strepdb.streptomyces.org.uk/cgi-bin/dc3.pl?accession=AL645882&serial=2769&width=900&start=3031362&end=3041362&iorm=map The second website below is the source of the DNA sequence we have used. http://www.ncbi.nlm.nih.gov/nuccore/NC_003888.3
This DNA sequence codes Lysine descarboxylase (Streptomyces coelicolor) is an enzyme from the lyase family that converts lysine to cadaverin. The enzyme realizes the carbonyl group of the lysin amino acid. There is cadeverin producing (1,5-diaminopentane), a primary diamine which alkaline environment. The lysine decarboxylase is an enzyme induced the synthesis of which is promoted by anaerobiosis and an acidic pH. In bacteriology, this enzyme is sought through the middle of Moeller lysine or medium lysine Taylor. This enzyme is also the first step in the production of desferrioxame B which is a siderophore.
The DNA sequence was modified by the addition of a prefix and suffix sequences (containing the restriction site EcoRI, XbaI and SpeI, PstI respectively) and was inserted in a promoteur using SLIC method.
Claire raynaud, Yoann Santin, Louis Givelet
The following website contains the gene reference and its chromosomal location. http://strepdb.streptomyces.org.uk/cgi-bin/dc3.pl?accession=AL645882&serial=2769&width=900&start=3031362&end=3041362&iorm=map Start : 3037106 End : 3038347 The coding sequence of DNA comes from http://www.ncbi.nlm.nih.gov/nuccore/NC_003888.3
This sequence codes Monooxygenase which is an enzyme that incorporate one hydroxyl group into substrates in many metabolic pathways. In this reaction, the two atoms of dioxygen are reduced to one hydroxyl and one H2O molecule by the concomitant oxidation of NAD(P)H. It is also the second step in the production of desferrioxamine B in Streptomyces coelicolor, allowing the transformation of cadeverine into N-hydroxycadaverine.
We had prefix and suffix to the DNA sequence containing restriction sites (EcoRI, Xba1 and SpeI, PstI respectively.
The following website gave us information about the gene reference and his chromosomal location. http://strepdb.streptomyces.org.uk/cgi-bin/dc3.pl?accession=AL645882&serial=2769&width=900&start=3031362&end=3041362&iorm=map Start : 3038344 End : 3038898 The coding sequence ised comes from the website below : http://www.ncbi.nlm.nih.gov/nuccore/NC_003888.3
desC is the coding sequence of DesC which is an acyl transferase of the Desferrioxamine production pathway. It is the enzyme of the third step that transform N-hydroxycadaverin into N-acetyl N-hydroxucadaverine by an Acetyl-CoA dependent manner.
We added prefix and suffix sequences containing the restriction sites (EcoR1, XbaI and SpeI, PstI respectively)
desD, desferrioxamine biosynthesis protein coding sequence
Gene reference and its chromosomal position have been found on the following website : http://strepdb.streptomyces.org.uk/cgi-bin/dc3.pl?accession=AL645882&serial=2769&width=900&start=3031362&end=3041362&iorm=map Start : 3038895 End : 3040682 The website bellows is the source of the DNA sequence we used. http://www.ncbi.nlm.nih.gov/nuccore/NC_003888.3
This coding sequence codes DesD od Streptomyces coelicolor, which is the last protein involved in the metabolic pathway of Desferrioxamine B. This enzyme is called desferrioxamine biosynthesis protein or DesD and allows to transform N-acetyl N-hydroxycadaverine into Desferrioxamine B by the transformation of 3a nucleoside triphosphate into 3a nucleoside 5'monophosphate-3-diphosphate.
We added prefix and suffix subsequences (containing EcoRI, XbaI and SpeI, PstI restriction sites respectively).
Conversion of the iGEM parts registry to SBOL2.1