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From 2010.igem.org

Molecular Mechanisms

Relevant properties of the chemotaxis pathway

Localization Che proteins

In [1] CheA, CheY and CheZ proteins were fused to YFP in E. coli mutants to determine their localization inside the cell. All three proteins tend to co-localize with methyl accepting chemotaxis proteins (MCPs, the receptor proteins of the chemotaxis pathway) at the membrane, but with different strengths.

• CheA and CheZ nearly only localize at the MCPs.
• CheY has a higher concentration around the MCPs, but is present in significant concentrations in the cytoplasm.

Consistent with common knowledge, CheA was not localized inside the cell in mutants lacking MCPs. CheZ showed no localization in ΔcheA mutants. CheY did not show any significant localization in ΔcheA mutants nor in mutants lacking a significant amount of receptor proteins.

Similar studies were done in [2] and [3] with a mutant expressing GFP-CheR. Around 50% of the cells showed a weak polar localization of the fusion proteins, whereas the other cells showed no localization at all.

Relevant properties of the PhyB/PIF3 module

Properties of holophytochrome

Phytochrome apoproteins (PHYs) are a family of 120-130kD soluble proteins [4, p.105]. They are covalently bound to a linear tetrapyrrole chromophore. The native holoproteins homodimerizes. It is likely that each subunit in the homodimer is phototransformed independently (no co-operative effects) [4, p. 131].

There are two mayor types of phytochromes, type I (PhyA) and type II (PhyB and PhyC) [4, p.142]. The Pfr form of type I proteins is labile (fast degradation, half live between 30min and 2h). Although the Pfr form of type II is also degraded faster than the corresponding Pr form, it is much more stable (half live around 8h). The degradation is temperature dependent and possibly an active process [4, p.142 ff]. On the other hand, the Pr form is very stable (half live of ofer 100h in Cucurbita).

In some plants pyhtochrome type I in the Pfr form sequesters [4, p.146]. The sequestering happens in the timescale of seconds, whereas even when ,deactivated and again and in the Pr form, the half life of the bulk is around 25 min at 25C for etiolated Avena. It is not clear if type II Pfr also sequesteres. It is also possible that the sequestering is part of the active degradation of type I Pfr [4].

Literature

[1] Sourjik and Berg: Localization of components of the chemotaxis machinery of Escheria coli using fluorescent protein fusions. Molecular Biology. 2000; 37:4.

[2] Shiomi, Banno, Homma, and Kawagishi: Stabilization of polar localization of a chemoreceptor via its covalent modifications and its communication with a different chemoreceptor. Journal of Bacteriology. 2005; 187:22.

[3] Shiomi, Zhulin, Homma, and Kawagishi: Dual recognition of the bacterial chemoreceptor by chemotaxis-specific domains of the CheR methyltransferase. The Journal of Biological Chemistry. 2002; 277:44.

[4] Kendrick and Kronenberg: Photomorphogenesis in plants. Kluwer academic publishers, Dordrecht, The Netherlands. 2nd edition, 1994.