Part table
| Basic parts | |
|---|---|
| Acetokinase | BBa_K3036001 |
| Phosphotransacetylase | BBa_K3036002 |
| rpoH P5 promoter | BBa_K3036003 |
| GFP with degradation conduction tag RepA | BBa_K3036006 |
| RelB with degradation-promoting tag RepA | BBa_K3036008 |
| Composite parts | |
|---|---|
| Cold triggered kill switch | BBa_K3036004 |
| Arabinose-induced suicide switch | BBa_K3036005 |
| Acetic acid overproducing pathway | BBa_K3036007 |
| Characterized parts | |
|---|---|
| RNA thermometer (FourU) | BBa_K115002 |
| RelE toxin | BBa_K185047 |
| RelB antitoxin | BBa_K185048 |
| pBAD strong | BBa_K206000 |
| MazF | BBa_K302033 |
| FadD | BBa_K861010 |
| FadE | BBa_K861020 |
BBa_K3036001, encodes acetokinase (ACK) of E. coli. The natural function of ACK is to convert acetate into acetyl phosphate. However, the reversibility of the reaction confers ACK a potential to produce acetic acid using acetyl phosphate as a precursor. Our team made use of this part, together with phosphotransacetylase (PTA), to overproduce acetic acid.
BBa_K3036002, encodes phosphotransacetylase (PTA) of E. coli. The natural function of ACK is to convert acetyl-CoA into acetyl phosphate, giving it a potential to enhance acetate production by providing its precursor. Our team made use of this part, together with acetokinase (ACK), to overproduce acetic acid.
BBa_K3036003, this rpoH P5 promoter is a glucose inhibitory promoter regulated by cAMP. This part is used as a digestion sensor to respond to glucose concentration changes, which is major digestive product of starch in small intestine. As a result, we can have our bilateral switch convert with the changing environment in human intestine.
BBa_K3036006, the RepA-GFP part is designed for increasing the degradation rate of green fluorescent protein (GFP) by adding a 16-amino-acid-long tag replication protein A (RepA) at the N-terminal, therefore the green fluorescence will degrade sooner when expression ends. Hence, when GFP is used as a reporter, it is more sensitive in monitoring the state of any inducible promoter.
BBa_K3036008, is RelB with degradation-promoting tag RepA. RelB is the inhibitor of toxin RelE. The two proteins can be made use of in a toxin-antitoxin system, where RelE gets constantly expressed and RelB is expressed only under certain conditions, validating a kill switch that is triggered when its host escapes to the outer environment.
BBa_K3036004, is kill switch consists of RNA thermometer (BBa_K115002), relE (BBa_K185000), and relB (BBa_K185048), aiming to prevent environmental contamination from the engineered bacteria through a cold-triggered toxin system.
BBa_K3036005, this arabinose-induced suicide switch consists of pBAD (BBa_K206000) and mazF (BBa_K302033), aiming to enable users of this microbe to terminate engineered bacteria inside their intestines with L-arabinose whenever needed. This suicide switch does no harm to human and can be used by direct in-taking of inducer.
BBa_K3036007, is an acetic acid overproducing pathway. The trait of PTA, together with the reversibility of the conversion catalyzed by ACK, gives them a potential to enhance acetate production when overexpressed. Moreover, we include fatty acyl-CoA synthetase (FadD), a key enzyme in beta-oxidation of higher fatty acids, to increase the yield of acetate to a further extend.
BBa_K115002, we characterize this RNA thermometer (FourU) part by an antitoxin-toxin system, in which the downstream gene encodes for a constantly expressed toxin (relE: BBa_K185000) and the upstream gene encodes for a labile antitoxin (relB: BBa_K185048) under the control of the RNA thermometer (FourU).
BBa_K185047, we characterize relE toxin by an antitoxin-toxin system, in which the downstream relE gene encodes for a stable toxin, and the upstream relB (BBa_K185048) gene encodes for a labile antitoxin under the control of a temperature-sensitive RNA thermometer (BBa_K115002).
BBa_K185048, we characterize relB by an antitoxin-toxin system, in which the downstream relE (BBa_K185000) gene encodes for a stable toxin, and the upstream relB gene encodes for a labile antitoxin under the control of a temperature-sensitive RNA thermometer (BBa_K115002).
BBa_K206000, we characterize pBAD by an induced suicide system, in which pBAD controls the downstream mazF (BBa_K302033) gene that serves as a reporter, which encodes an endoribonuclease that cleaves RNAs at ACA sites and causes the death of microbe. As a result, we can characterize pBAD in a cell density-dependent manner in Escherichia coli K-12.
BBa_K302033, we characterize mazF by an induced suicide system, in which the downstream mazF (BBa_K302033) gene encoding toxin MazF is put under the control of L-arabinose induced promoter pBAD (BBa_K206000). MazF is an endoribonuclease that cleaves RNAs at ACA sites and causes the death of microbe as a reporter. As a result, we can characterize mazF in a cell density-dependent manner in Escherichia coli K-12.
BBa_K861010, we overexpress fadD gene derived from E. coli K-12 DH5alpha genome in our engineered intestinal microbe to promote degradation of higher fatty acids which would otherwise be assimilated by human body.
BBa_K861020, we overexpress fadE gene derived from E. coli K-12 DH5alpha genome in our engineered intestinal microbe to catalyze dehydrogenation process from fatty acyl-CoA to fatty enoyl-CoA in fatty acid beta oxidation.