Team:Washington/Alkanes/Future

From 2011.igem.org

Our current in vivo alkane production system efficiently makes C15 alkanes. However, to be efficient enough for factory production, there are two broad goals to be done:

1. Increase production efficiency
2. Increase the diversity of the range of alkanes that can be produced.
3. Increase the scale of the system for industrial processes.

We have already begun efforts to expand the efficiency and scope of alkane production, check them out below!

System Optimization

The efficiency of this system was reported to be much higher than our initial system, but the growth conditions of the assay and the DNA was not available to us. We increased production efficiency by altering the initial environmental conditions in the assay.

Decarbonylase Redesign

One way to diversify the kind of alkanes produced is to alter the substrate specificity of the proteins involved. Since an alternative alkane-producing enzyme has not been found, we decided to mutate the aldehyde decarbonylase to produce shorter-chain alkanes.

Alternative Aldehyde

Another way to diversify our system is to use alternative proteins. Our current system uses acyl-ACP reductase, and we've identified an hypothetical alternative system that produces aldehydes: LuxCDE, made from parts from the 2010 competition.

Branched Alkane Biosynthesis

Our system is only capable of producing unbranched alkanes, as the cell mainly utilizes straight chained fatty acids. However, fuel we use are also composed largely of branched alkanes that affect very important properties of the fuel such as flash point and freezing point. If our fuels are truly intended to be synthesized in bacteria, we need to work on methods of making those crucial branched chained alkanes. We explored FabH2, a protein that when involved in fatty acid synthesis makes branched fatty acids.

Enzyme Localization

The easiest way to increase yield is to increase the concentration; if the number of molecules remained the same but the volume decreased, the reaction speed increases with occurrences of molecular collisions. However, this is only easily done with purified protein assays. However, there are ways to "increase the concentration" of reactions in cells. We cannot make the cells literally denser, but we can localize the enzymes involved in the reaction, which either decreases the volume of the cell in which the enzymes reside and/or limit the number of reaction steps.

Alternative Chassis

By synthesizing alkanes in organisms, alkanes can become a useful renewable energy source; CO2 emitted from burning the alkanes are converted to glucose through photosynthesis, and then the glucose can be fed to alkane-synthesizing organisms to restart the cycle. This cycle can be drastically improved if there were fewer steps in the cycle. We explored the possibility of optimizing the alkane biosynthesis system in different micro-organisms, with the idea that perhaps with autotrophic organisms, we do not need to obtain glucose for the reaction.

System Modeling

We have begun Flux Balance Modeling the alkane production system in order to better understand the carbon flux through the pathway. We believe that by understanding the curent flux through the pathway we can intelligently reengineer the platform in order to maximize flux to alkanes.