Part:BBa_K2622006
Mistic (Mstx)
Mistic (Mstx) is an unique hydrophilic protein from Bacillus subtilis that works as a fusion tag and helps to integrate other proteins into lipid membranes more easily. When fused to the N-terminus of integral membrane proteins, MstX enables the cargo proteins to fold into their native conformations in the membrane, thus yielding a high-level expression. Therefore, this biobrick has RFC12 prefix and suffix that allows to fuse proteins to the N or C-terminus in a stop codon free way.
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]
Contribution: iGEM22_WHU-China
Mistic possesses two features whose relationship has remained unclear: its interaction with the lipid bilayer and its ability to promote the overexpression of heterologous membrane proteins. The results of the article reveal that these two features are not correlated, as they could express the Y4 receptor, a human GPCR, also when fused to only H3 from Mistic.
Fig.1 (A) Comparison of maps of vectors pDXW-1, pDXW-2, pDXW-3 and pDXW-3-LRaceE. (B)Construction of the C. glutamicum aceE mutant strain YTW-1. (C and D) Growth difference ofC. glutamicum ATCC13032 and YTW-1 on CGXII media in the presence (C) or absence (D) of sodium acetate.
Results suggest that the presence of N-terminal sequences at the mRNA level rather than the ability of the translated product to interact with the membrane is important for obtaining high expression levels of Mistic-tagged protein constructs.
The production and, consequently, the localization in the inner membrane of GPCRs and other membrane proteins can be promoted by using N-terminal soluble E. coli proteins such as the glutathione S-transferase (GST)24 or the more recently reported YbeL and YnaI. The artical concluded that the absence of defined mRNA structures at the initiation region has a strong effect by facilitating translation initiation.
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