Difference between revisions of "Part:BBa K590034"

Revision as of 17:25, 26 October 2011

FabH2

This part encodes FabH2. [http://2011.igem.org/Team:Washington 2011 University of Washington iGEM Team] has produced even chain length alkanes using this part and the Petrobrick. In addition, expression of this part and the Petrobrick should theoretically produce branched chain alkanes, but we have not been able to demonstrate this effect, possibly due to the absence of the appropriate substrates in E. coli

Usage and Biology

FabH2 is from Bacillus subtilis. The FabH family of proteins initiates fatty acid elongation by converting an Acyl-CoA into an Acyl-ACP, with is extended by 2 carbon units to form longer chain length fatty acids. Normally, FabH proteins use a simple 2-carbon acetyl-CoA to start fatty acid biosynthesis, resulting in linear fatty acids. However, FabH2 can also use Isobutyryl-CoA, Isovaleryl-CoA, and 2-Methylbutyryl-CoA (products from Valine, Leucine, and Isoleucine degradation), resulting in 2-methyl branched fatty acid production. In addition, FabH2 has been hypothesized to start fatty acid elongation with a straight 3-carbon unit(propionyl-CoA), yielding odd chain length fatty acids, which could be converted into even chain length alkanes by thePetrobrick. Expression of FabH2 on the same high copy number constitutive plasmid as the PetroBrick( as in Part BBa_K590030) results in slow cell growth( insert picture), and low alkane yield( under 10 mg/L vs. approximately170 mg/L in the Petrobrick). [[Image:Washington_growthcurve.png|center|450px|thumb|Growth curve showing that cells expressing FabH2 are growth deficient.[ADC AAR DrR FabH2 ]]

By moving FabH2 into a low copy number inducible vector(As in The FabBrick), we were able to get C14 and C16 alkane production, completing the alkane spectrum from C13 to C17. This is the first time that even chain length alkanes have been recombinately produced.

GCMS trace confirming C16 alkane produced only upon FabBrick induction.

MS spectrum verifies peak contains C16 alkane.

Sequence and Features