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Self-assembly is important for TIP47 function in mannose 6-phosphate receptor transport
Paul M Sincock1, Ian G Ganley, Jeffrey P Krise
1Department of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305-5307, USA.
Traffic (Copenhagen, Denmark)
|January 22, 2003
Summary
Tail-interacting protein of 47 kDa (TIP47) binds mannose 6-phosphate receptors. Its N-terminal domain drives TIP47 oligomerization, essential for receptor transport but not binding.
Area of Science:
- Cell Biology
- Molecular Biology
- Protein Trafficking
Background:
- Mannose 6-phosphate receptors (MPRs) mediate lysosomal enzyme targeting.
- Tail-interacting protein of 47 kDa (TIP47) interacts with MPR cytoplasmic domains.
- TIP47 is implicated in MPR transport from endosomes to the trans-Golgi network.
Purpose of the Study:
- To investigate the role of TIP47 oligomerization in MPR binding and transport.
- To identify the domain responsible for TIP47 oligomerization.
- To elucidate the functional significance of TIP47 oligomerization in cellular trafficking.
Main Methods:
- In vitro binding assays using recombinant TIP47 fragments.
- Gel filtration and chemical cross-linking to assess TIP47 oligomeric state.
- Mammalian cell expression systems to study protein interactions and aggregation.
- In vivo transport assays to evaluate MPR trafficking.
Main Results:
- TIP47 exists as a homo-oligomer (likely hexamer) in the cytosol.
- An N-terminal domain (residues 1-151) is responsible for TIP47 oligomerization.
- TIP47 oligomerization is not required for binding to MPRs.
- TIP47 oligomerization is essential for stimulating MPR transport in vivo.
Conclusions:
- TIP47 possesses an N-terminal oligomerization domain.
- Oligomerization is separable from MPR binding function.
- TIP47 oligomerization is a critical regulatory step for MPR-mediated protein transport.