hahn lab

published molecular tools by protein target

Genetically encoded biosensors
Different designs with different advantages.

  • Cdc42
  • Rac1
  • RhoA
  • RhoC

Dye-based biosensors
Currently requires microinjection, electroporation etc.

  • Calmodulin
  • Src family proteins

Photomanipulation of proteins:  analogs of the target protein that are inhibited or activated by light
Precise control of both timing and localization.
light activated:

  • Rac1
  • Cdc42 (a required mutation in the effector binding site may affect effector interactions)
light-activated dominant negative:
  • Rac1

Photomanipulation of proteins: embed inhibitory or activating peptides or protein fragments in a protein construct such that they are exposed upon irradiation.
This controls endogenous protein.

  • PKA inhibition
  • MLCK inhibition
  • formin activating peptides (in collaboration with Zaidel-Bar group)

Engineered allosteric activation - proteins are activated upon addition of rapamycin (or a non-immunosuppressive analog) to the cell medium
Embed an engineered domain into a protein, rendering the active site inert until a membrane-permeable small molecule  is added to the cell medium. We have a general approach to apply this to other kinases as well as other protein families (unpublished).

  • Src
  • Fyn
  • Yes
  • Lyn
  • FAK
  • p38

Upon activation by rapamycin, a kinase can only interact with one specific downsream target
When the kinase is activated by rapamycin, it will interact specifically with only one of its multiple targets

  • Src activated and restricted to interaction with p130Cas
  • Src activated and restricted to interaction with FAK

Photomanipulation of protein activity: Control the rapamycin-mediated methods above with light
We have used a 'caged' analog of rapamycin, an analog whose activity is blocked by a photocleavable protecting group, to confer light control on the rapamycin-mediated methods above.

Photo-control of nuclear translocation (with Kuhlman group)
Control the exposure of nuclear localization and nuclear export sequences, thus regulating nuclear localization with light.

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