Part:BBa_K3634002

ATP-Grasp (ATPG) (Codon Optimised for E.coli)

Codon optimised adenosine triphosphate (ATP)-grasp (ATPG) CDS for use in E.coli as part of the shinorine synthesis gene cluster. The original gene for ATPG (Ava_3856) can be taken from the cyanobacteria species Anabaena variabilis ATCC 29413. The enzyme catalyses the conversion of 4-deoxygadusol (4-DG), produced by O-MT in the previous step of the reaction, to the mycosporine-like amino acid (MAA) mycosporine glycine. ATPG is proposed to phosphorylate 4-DG before conjugate addition occurs at the cyclohexanone ring by the nitrogen of glycine.

This part has been previously categorised by the Minnesota iGEM team of 2012 who also sought to express the enzyme in E.coli (BBa_K814001). Here, by using the IDT codon optimisation tool, we have optimised the GC content of the sequence taken from A.variabilis to improve production efficiency of the final product shinorine. We have further made the part biobrick assembly standard RFC[10] & RFC[1000] compatible by removing EcoR1 and PstI restriction sites introduced by this optimisation step.

The part should be used alongside the additional optimised parts (BBa_K3634000, BBa_K3634001 and BBa_K3634003) responsible for the ultimate conversion of the substrate sedoheptulose 7-phosphate to the final product shinorine. Shinorine is a UV absorbing compound which absorbs light of wavelength 333nm. It is produced by aquatic bacteria and algae species as a source of protection in regions of high UV intensity. Shinorine is just one of a whole selection of MAAs which provide UV photoprotection across a range of different wavelengths in the UV portion of the EM spectrum.

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
  COMPATIBLE WITH RFC[1000]