While working on project “RECOVER“, our team added the upcoming new basic and composite parts to the iGEM registry. The CO₂ fixation and the production of isobutanol is facilitated by the following part sequences.

Phosphoribulokinase is responsible for converting ribulose-5-phosphate into ribulose-1,5-bisphosphate, a vital substrate for CO₂ fixation by the RuBisCO enzyme. This conversion is a key step in the Calvin cycle, essential for photosynthesis and the production of organic compounds.

Phosphoribulokinase and Rubisco are co-engineered on a single plasmid to facilitate the CO ₂ fixation phase. This process ultimately yields 3-phosphoglycerate as the end product. Subsequently, 3PG is transformed into pyruvate, which then enters the ketoacid pathway, culminating in the production of isobutanol.

Boosts isobutanol production, genetically enhancing E. coli by increasing AdhA enzyme levels, compensating for the lack of KivD, and improving our biofuel manufacturing process.

Ribulose-1,5-bisphosphate carboxylase/oxygenase, a pivotal biological enzyme, occupies a central position in the process of carbon fixation. It strongly promotes the transformation of ribulose-1,5-bisphosphate, and carbon dioxide into two crucial molecules of 3-phosphoglycerate, which serve as indispensable building blocks for carbohydrate synthesis.

The subunit RbcX of the RUBISCO protein structure is an integral part of the protein. While it plays no part in the protein folding process [1].
It is present in both the cytoplasm as well as within a carboxysome. Therefore, for extraction, a simple crude cell lysis will be sufficient [2].
It is so integral that delayed post translational addition of RbcX resulted in the rapid loss of RbcL assembly and in the formation of insoluble RbcL. This indicates that RbcX has a specific chaperone function in preventing RbcL misassembly downstream of chaperonin [3].
The subunit RbcX of the RUBISCO protein structure is an integral part of the protein. While it plays no part in the protein folding process [1]. It is present in both the cytoplasm as well as within a carboxysome. Therefore, for extraction, a simple crude cell lysis will be sufficient [2]. It is so integral that delayed post translational addition of RbcX resulted in the rapid loss of RbcL assembly and in the formation of insoluble RbcL. This indicates that RbcX has a specific chaperone function in preventing RbcL misassembly downstream of chaperon n [3]. To ensure this delay in transcription does not occur, it is suggested that a strong promoter is added ups ream of the RbcX coding sequence to prevent any delays in translation and thereby prevent any missassembly of the other RbcL subunits. We have added this information for our existing part BBa_K4689060 to make part usage easier.

Many teams including ours would have a branching point in their metabolic cycle due to a new engineered pathway in their organism for a substrate already present in the organism. This means there is increased demand for an existing substrate in a limited concentration. Unfortunately there is no widely used enzyme kinetics model present for this competition as it is present for substrate inhibition [such as competitive or in-competitive models of inhibition where multiple substrates or substrate like analogs are present]. To solve this, our team conducted literature research and found an unstudied model of enzyme kinetics where two enzymes are competing for a single substrate.[4]

Our team has also modeled the metabolic pathway branchpoint in our enzyme kinetics using this formula. For this purpose, it was uploaded as a user defined function on COPASI.