Part:BBa_K2675000
AimR of phage phi3T
This part is an Escherichia coli K12 codon optimized version of AimR, the receptor of the arbitrium peptide of phage phi3T [1].
It is an improvement of BBa_K2279000.
Usage and Biology
The switch from lytic-to-lysogenic cycle of the Bacillus phage phi3T is based on the expression of a single transcript, AimX [1]. The expression of AimX is controlled by AimR who was described to be a transcriptional regulator of the pAimX promoter in B. subtilis [1].
The AimR activity is modulated by the interaction with a hexapeptide (SAIRGA in phage phi3T). Binding of SAIRGA to AimR blocks the activator function of AimR that is no longer able to bind to the pAimX promoter and consequently, the AimX gene is repressed. This facilitates a switch from lytic-to-lysogenic viral cycle. By acting as a negative regulator of AimR, the SAIRGA signal makes the lysis-to-lysogeny switch of phi3T phage dependent on the “quorum” of phi3T phages in the bacterial population.
From a structural point of view, AimR shows a dimeric arrangement of 88 kDa, and each subunit has 378 amino acids and a molecular weight of 44 kDa. Each monomer consists of 20 α-helices, eight of which assemble into four atypical TPR motifs that form the peptide binding pocket together with other helices [2, 3]. This topology makes AimR structurally related to the RNPP family of quorum sensing systems of Gram-positive bacteria. The interaction between the AimR monomers is mediated by a C-terminal capping helix that was interacting with its counterpart on the other monomer and is mainly mediated by Van Der Waals interactions [3]. The peptide binding causes a big structural change and even dissociation into monomers for AimR and is preventing the DNA binding activity of AimR.
To express this AimR in E. coli, the sequence was codon optimized for E. coli DH5α, a specific RBSs (BBa_K2675010) was designed with the Salis RBS Calculator [4, 5] and the peptide was placed under the control of the constitutive strong promoter (BBa_J23102). Thus, the composite part BBa_K2675040 was generated and proved to be functional in E. coli: it expresses AimR that behaves as a transcriptional regulator of sfGFP-LVAtag expression from BBa_K2675058 and BBa_K2675059.
References
[1] Erez Z, Steinberger-Levy I, Shamir M, Doron S, Stokar-Avihail A, Peleg Y, Melamed S, Leavitt A, Savidor A, Albeck S, Amitai G, Sorek R. Communication between viruses guides lysis-lysogeny decisions. Nature (2017) 541, 488-493.
[2] Dou C, Xiong J, Gu Y, Yin K, Wang J, Hu Y, Zhou D, Fu X, Qi S, Zhu X, Yao S, Xu H, Nie C, Liang Z, Yang S, Wei Y, Cheng W. Structural and functional insights into the regulation of the lysis-lysogeny decision in viral communities. Nat Microbiol (2018) doi: 10.1038/s41564-018-0259-7.
[3] Wang Q, Guan Z, Pei K, Wang J, Liu Z, Yin P, Peng D, Zou T. Structural basis of the arbitrium peptide-AimR communication system in the phage lysis-lysogeny decision. Nat Microbiol (2018) doi: 10.1038/s41564-018-0239-y.
[4] Espah Borujeni A, Channarasappa AS, Salis HM. Translation rate is controlled by coupled trade-offs between site accessibility, selective RNA unfolding and sliding at upstream standby sites. Nucleic Acids Res (2014) 42, 2646-2659.
[5] Salis HM, Mirsky EA, Voigt CA. Automated design of synthetic ribosome binding sites to control protein expression. Nat Biotechnol (2009) 27, 946-50.
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]
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