Team:Baltimore Bio-Crew/Composite Part

Baltimore Bio-Crew


Bio-Engineering E.Coli To Degrade Plastic and Save The Baltimore Inner Harbor

Composite Parts

We designed two constructs, known as biobricks, to allow the bacteria to express the proteins lipase and esterase that will degrade PET plastics. These constructs are identical except for the sequence for the enzyme each construct would produce: esterase and lipase. The chlorogenate esterase construct (BBa_K2436001) was cloned successfully and submitted to the registry. The lipase construct was not cloned successfully. Our constructs were designed as follows.
Lactose Regulated Promoter (BBa_R0011): a DNA sequence that starts the transcription process in the presence of lactose or IPTG. Thus, the promoter is only active in certain times to regulate the amount of proteins the bacteria is able to produce at a time. The promoter causes the gene to activate and begin the process of forming mRNA and eventually proteins.
Ribosome Binding Site (BBa_B0030): an RNA sequence found in mRNA to which ribosomes can bind and initiate translation. It is a sequence of nucleotides upstream of the start codon of an mRNA transcript that is responsible for the recruitment of a ribosome during the initiation of protein translation.
PelB Secretion Tag (BBa_K2010002): allows for bacteria to secrete the enzymes (lipase and esterase) into the periplasm. Once the enzymes are secreted to the periplasm, they are able to exit the cell on their own.
Protein Coding Sequence (BBa_K2187001 for esterase or BBa_K2187000 for lipase): the DNA sequence is the lipase and esterase gene sequences. The lipase and esterase are both needed to successfully produce the plastic degrading enzymes because both work together to break down and eliminate the plastic particles. One construct contains the chlorogenate esterase coding sequence, and the other construct has the lipase coding sequence.
His Tag: allows us to purify the protein on a nickel column once it’s secreted outside of the bacteria
Terminator (BBa_K864600): a sequence that occurs at the end of the gene and causes transcription to stop