We are considering two different methods of implementation. The first, would be to engineer an Homogalacturonan (HG) degradation pathway into a microbe, lyse the cells after protein expression of the degredation pathway occurs, then add its lysate to the compost. Secondly, we could purify each of our enzymes and create a lyophilized powder that can be added into the compost. To test our system, one assay we could do is a colourimetric assay to test the degradation properties of the enzymes, by measuring the amount of galactose present when they break down HG.
The first potential host we have considered which we would transform to express our desired enzymes is E.coli, as it’s presently the best understood and most widely used organism in synthetic biology; with rapid growth and high protein yields, it provides a model organism to study bacterial metabolic pathways. Additionally, the genome does not have any pectin-degrading enzymes, a great candidate to characterize and validate our HG degradation system.
Another organism we have considered is Bacillus Subtilis, which is classified as safe, and not considered pathogenic or endotoxic to humans, as it’s even used in some probiotic supplements. Additionally, some strains are able to utilize polysaccharides such as pectin as the carbon source, so pectinases may be present in its genome. This could be either helpful or problematic to our system. On the one hand, its pectinases could help improve the efficiency of our system. On the other, this may make it hard to validate and characterize our proposed system.
Our inspiration in making a product that addresses food waste problems is the fact that our hometown Lethbridge Alberta has no curbside or drop off compost options and the weather inhibits many from doing their own composting, and this results in food waste making up nearly half of all Lethbridge landfill waste. Our project tpectinACE is intended to be used in personal composts in any climate. We are hoping by increasing the public interest in composting, the city of Lethbridge will start a “green cart program” like much of Canada has already. Instead of relying on at home composting, an infrastructure could be built in our community to take care of household compost.
tPectinACE comes with its disadvantages if not used correctly or in the correct way. We are going to use enzymes like PelB or PelC to degrade Homogalacturonan protein which have sections that are still unidentifiable and so if the right temperature is not applied(50 - 158 degree Celsius) the decomposition process might not have the desired effects and be as efficient. Once in use, the composter should be kept away from small children and pets because the products used have not been tested on the human body and the effects can cause disruption to the systems of the body and its processes. Our system may be available in a purified enzyme form or as a GMO lysate. Either way, the system must not be ingested or inhaled or handled by little kids, pets or anyone who has health issues for reasons unknown. The suggested amount should be applied as instructed (based on our future experiments) because too little could lead to an unsatisfactory compost or one that will not have the desired effect which could lead to heavy odour and an unstable environment for organisms to survive in. But if all instructions are followed, our system, tPectinAce, will improve composting and make it easier for everyone and will ensure a safe environment that has reduced waste and increased recycling.