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From 2011.igem.org

Parts

The 2011 University of Glasgow team has submitted 9 novel biobricks. You can find out more about these below, or by visiting the Registry Page. If you are interested in finding out about the safety reviews of these biobricks then please visit our Biobrick Safety Page, which can be accessed here. You can also view links to further information on the Biobrick Safety page.

Phosphodiesterase

Phosphodiesterases are a family of enzymes naturally present in microorganisms, which break phosphodiester bonds. The specific enzyme we are using breaks the phosphodiester bond in the second messenger nucleotide, cyclic digaunylate (cyclic-di-GMP). Cyclic-di-GMP is important in many bacterial processes, including biofillm formation and motility. The enzyme regulates signal transduction by controlling levels of the signalling molecule in cells.

This enzyme was amplified from the genome of Pseudomonas aeruginosa PA01. If you want to view specific information on this enzyme, please click here.

We are using the enzyme to control the levels of cyclic-di-GMP within the cell. We expect that the targeted expression of phosphodesiterase could be used to interfere with biofilm formation or to trigger dispersal.

Due to the number of processes in which cyclic-di-GMP is used by prokarytes, we hope that future iGEM teams will find diverse uses for it.

Phosphodiesterase with 6xHis Tag

We have also created a version of the phosphodiesterase biobrick with a 6xHIS tag for affinity purification.

Novel Reporters

We have submitted two novel reporters to the registry, LOV2 and iLOV. Read on to find out about the exciting advantages these reporters have over GFP derived flourescent proteins.

LOV2

The LOV (Light-Oxygen-Voltage) domain is a photoreceptor that responds to blue light. In nature it was first found to be involved in the phototropism response in plants and has since been found to be present in fungi and bacteria also. It has been shown to be coupled to many domains, for example phosphodiesterase or kinases.

We are using it as a reporter due to its ability to function in anoxic conditions. This is particularly useful in biofilms and is a function that fluorescent proteins derived from GFP do not have.

LOV2 with 6xHIS tag

We have also created a version of the LOV2 biobrick with a 6xHIS tag for affinity purification.

iLOV

iLOV is a version of LOV2 that has been altered through site directed mutagenesis and DNA shuffling. iLOV has the same function and uses as LOV2, but contains mutations that caused increased intensity in brightness.

As a reporter, it is advantageous over GFP derived fluorescent proteins due to its small size (useful if you are tagging proteins), ability to recover quickly from photobleaching and use in anoxic conditions.

Surfactant Proteins

We have developed biobricks of surfactant proteins, Ranaspumin and Latherin, which have been investigated for their antimicrobial activity.

Ranaspumin (RSN2)

Ranaspumin is a protein found in the foam nests of the Túngara Frog (Engystomops pustulosus). In nature it is used to protect and incubate the fertilized eggs of the frog.

It has natural antimicrobial and antibiofilm activity.

Ranaspumin is a particulary interesting protein due to its unique amino acid sequence.

Ranspumin (RSN2) with 6xHIS tag

We have also created a version of Ranaspumin (RSN2) biobrick with a 6xHIS tag for affinity purification.

Latherin

Latherin is a surfactant protein that was originally isolated from horse sweat. Its normal biological function is temperature regulation and is believed to function by enhancing evaporation from the pelt. Due to its ability to bind to hydrophobic surfaces, the protein is being investigated for its possible ability to aid in breaking up biofilms.

We are using latherin to aid in dispersal of biofilms, which is a natural property of the protein.

Latherin with 6xHIS tag

We have also created a version of Latherin with a 6xHIS tag for affinity purification.