A composite part is a functional unit of DNA consisting of two or more basic parts assembled together. BBa_I13507 is an example of a composite part, consisting of an RBS, a protein coding region for a red fluorescent protein, and a terminator. New composite BioBrick devices can be made by combining existing BioBrick Parts (like Inverters, Amplifiers, Smell Generators, Protein Balloon Generators, Senders, Receivers, Actuators, and so on).

GlgCAB for polycistronic expression of glycogen synthesis genes in E.coli

This composite part combines all 3 enzymes involved in glycogen synthesis. The ADP-glucose pyrophophorylase (GlgC) forms ADP-glucose from ATP and glucose-1-phosphate, the glycogen synthase (GlgA) elongates alpha-1,4-linked chains and the branching enzyme (GlgB) catalyzes the formation of alpha-1,6-linked branches. This construct is an extension and improvement of Part:BBa_K118016.

Application

In order to upregulate the whole glycogen synthesis pathway, the polycistronic plasmid was built. The glgC coding sequence is based on the part BBa_K118016 from Team Edinburgh 2008, but he RBS B0034 was added to the existing Biobrick. The construct was confirmed by sequencing. The expression of all three enzymes GlgC, GlgA and GlgB was tested in Bl21 Gold (DE3) strains containing BBa_K1585321 in a pSB1A30 expression vector.

SDS-PAGE of glgCAB in pSB1A30 GlgA was expressed in pSB1K30 and compared to mRFP expression. The small arrows point to the GlgA bands of 52.4 kDa. No difference between induced and uninduced was observed. After overnight expression the strong bands were still present.