Coding
PhyB-900

Part:BBa_K365002

Designed by: LACOTTE Yohann   Group: iGEM10_ESBS-Strasbourg   (2010-09-14)

Phytochrome B from A. thaliana (aa 1-908)

Note: This is a N-part RFC[25] Biobrick which means only protein fusions to the C-terminus of this Biobrick is possible. For a improved version of this Biobrick which also allow fusion to the N-terminus of PhyB (e. g. Strep-tag II or His-tag), please see here: BBa_K801031

This part consists in the first 908 amino-acids of Phytochrome B of A. thaliana. When bound to its natural chromophore Phycocyanobiline (PCB) it can reach its Pfr activated state and physically interact with different PIF.

Background:

Phytochromes characterised by a red/far-red photochromicity. Through red-light (650–670 nm) absorption the phytochrome undergoes a rapid conformational change from its ground state Pr to its active state Pfr. The structural change allows the binding of the PIF. This light-sensitive interaction has been mapped to the 650-residue amino-terminal photosensory core of PhyB (Khanna et al., 2004). The process is completely reversible through absorption in the near infra-red spectrum (705–740 nm).

The photoreceptor protein PhyB serves for the light-dependent activation of the system, therefore it will be fused to the N-teminal of the ClpX-trimer.


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Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 490
    Illegal BamHI site found at 572
    Illegal XhoI site found at 523
    Illegal XhoI site found at 542
    Illegal XhoI site found at 2677
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2062
    Illegal BsaI site found at 2468
    Illegal SapI site found at 739


Conception:

In in-vivo applications it has been shown that the PIF-interaction with the PhyB photosensory core (residues 1–650) is irreversible in infrared light. Lim & Voigt (2009) demonstrated by assaying PIF6 (which has the strongest interactions of all previously reported PIF domains) against different variants of PhyB that the tandem C-terminal PAS domains (residues 1-908)of plant phytochromes are necessary to confer rapid photoreversibility under infrared light. The original sequence contains a SpeI restriction within the first 908 residues.

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