Difference between revisions of "Part:BBa K2271060"
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− | [[File:Snc1duscgn.png|200px|thumb|right|'''Figure 1.''' A diagram of the general domain structure of Snc1. V is a variable domain which is not important for the binding to the t-SNARE. TM is the transmembrane domain. H1 and H2 are the α-helical segments, forming the SNAREpin with the t-SNARE | + | [[File:Snc1duscgn.png|200px|thumb|right|'''Figure 1.''' A diagram of the general domain structure of Snc1. V is a variable domain which is not important for the binding to the t-SNARE. TM is the transmembrane domain. H1 and H2 are the α-helical segments, forming the SNAREpin with the t-SNARE. [https://parts.igem.org/Part:BBa_K2271060:Design [1]]]] |
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− | This part is a truncated version of the v-SNARE ('''v'''esicle- synaptosome-associated-'''S'''oluble '''N'''-ethylmaleimide-sensitive-factor '''A'''ttachment '''RE'''ceptorprotein) Snc1. Snc1 is in the wildtype form involved in the fusion of Golgi-derived secretory vesicles with the plasma membrane. Domains of the protein are a variable domain which is not important for the binding to the t-SNARE, H1 and H2 are the α-helical segments (forming the SNAREpin with the t-SNARE) and the transmembrane domain [https://parts.igem.org/Part:BBa_K2271060:Design | + | This part is a truncated version of the v-SNARE ('''v'''esicle- synaptosome-associated-'''S'''oluble '''N'''-ethylmaleimide-sensitive-factor '''A'''ttachment '''RE'''ceptorprotein) Snc1. Snc1 is in the wildtype form involved in the fusion of Golgi-derived secretory vesicles with the plasma membrane. Domains of the protein are a variable domain which is not important for the binding to the t-SNARE, H1 and H2 are the α-helical segments (forming the SNAREpin with the t-SNARE) and the transmembrane domain.[https://parts.igem.org/Part:BBa_K2271060:Design [1]] |
Revision as of 22:51, 30 October 2017
Snc1
Usage and Biology
]This part is a truncated version of the v-SNARE (vesicle- synaptosome-associated-Soluble N-ethylmaleimide-sensitive-factor Attachment REceptorprotein) Snc1. Snc1 is in the wildtype form involved in the fusion of Golgi-derived secretory vesicles with the plasma membrane. Domains of the protein are a variable domain which is not important for the binding to the t-SNARE, H1 and H2 are the α-helical segments (forming the SNAREpin with the t-SNARE) and the transmembrane domain.[1] For our approaches we used a truncated version without the transmembrane domain. This Snc1 truncation was fused to the N-Terminus of different peroxisomal membrane anchor (Pex15 /PEX26) to secrete the compounds of this compartment.
Experimental design
For testing this part we used a fusion with the N-terminus of a peroxisomal membrane anchor. We co-expressed this construct with GUS-PTS1 (beta-glucuronidase)to perform a [http://2017.igem.org/Team:Cologne-Duesseldorf/Experiments GUS Assay]. The secreted GUS in the supernatant was measured with the turnover of 4-methylumbelliferyl-beta-D-glucuronide to 4-methyl umbelliferone (4-MU). The fluorescent 4-MU was measured with a plate reader (excitation: 365 nm, emission: 465 nm).
Results
Different peroxisomal membrane anchors were tested using the GUS-assay. The highest activity of GUS could be measured in the supernatant of Pex15 as a membrane anchor.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]