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Lee Pullan,
Srinivas Mullapudi,
Zhong Huang,
Philip R Baldwin, Christopher Chin,
Wei Sun,
Susan Tsujimoto,
Steven J Kolodziej,
James K Stoops,
J Ching Lee,
M Neal Waxham,
Andrew J Bean,
Pawel A Penczek
[show abstract]
[hide abstract]
ABSTRACT: The structure of the endosomal-associated protein, Hrs, has been determined with cryo-electron micros-copy. Hrs interacts with a number of proteins, includ-ing SNAP-25 and STAM1, forming a complex that binds ubiquitin moieties. Analytical ultracentrifugation stud-ies revealed that Hrs exists as a hexamer. The symme-try and the structure of the hexameric form of Hrs were determined with the single-particle reconstruction method. Hrs comprises three antiparallel dimers with a central core and distinct caps on either end. Crystal structures of VHS and FYVE domains fit into the Hrs end caps in the EM density map. Thus, the location of domains that interact with the endosomal membrane, the VHS, FYVE, and C-terminal domains, facilitates the anchorage of Hrs to the membrane, initiating the functional processes of Hrs on the endosome. Based on our model, the Hrs hexamer interacts with the mem-brane and acts as a ''master molecule'' that presents multiple sites for protein binding.
Structure 05/2006; 14:661-671. · 6.35 Impact Factor
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Lee Pullan,
Srinivas Mullapudi,
Zhong Huang,
Philip R Baldwin, Christopher Chin,
Wei Sun,
Susan Tsujimoto,
Steven J Kolodziej,
James K Stoops,
J Ching Lee,
M Neal Waxham,
Andrew J Bean,
Pawel A Penczek
[show abstract]
[hide abstract]
ABSTRACT: The structure of the endosomal-associated protein, Hrs, has been determined with cryo-electron microscopy. Hrs interacts with a number of proteins, including SNAP-25 and STAM1, forming a complex that binds ubiquitin moieties. Analytical ultracentrifugation studies revealed that Hrs exists as a hexamer. The symmetry and the structure of the hexameric form of Hrs were determined with the single-particle reconstruction method. Hrs comprises three antiparallel dimers with a central core and distinct caps on either end. Crystal structures of VHS and FYVE domains fit into the Hrs end caps in the EM density map. Thus, the location of domains that interact with the endosomal membrane, the VHS, FYVE, and C-terminal domains, facilitates the anchorage of Hrs to the membrane, initiating the functional processes of Hrs on the endosome. Based on our model, the Hrs hexamer interacts with the membrane and acts as a "master molecule" that presents multiple sites for protein binding.
Structure 05/2006; 14(4):661-71. · 6.35 Impact Factor
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[show abstract]
[hide abstract]
ABSTRACT: Chemokines play a fundamental role in trafficking of immune cells and in host defense against infection. The role of chemokines in the recruitment process is highly regulated spatially and temporally and involves interactions with G protein-coupled receptors and cell surface glycosaminoglycans. The dynamic equilibrium between chemokine monomers and dimers, both free in solution and in cell surface-bound forms, regulates different components of recruitment such as chemotaxis and receptor signaling. The binding and activity of the chemokine interleukin-8 (IL-8) for its receptors, previously studied using "trapped" non-associating monomers and non-dissociating dimers, show that the monomer has a native-like function but support conflicting roles for the dimer. We have measured the binding of native IL-8 to the CXCR1 N-domain, using isothermal titration calorimetry and sedimentation equilibrium techniques. The N-domain constitutes a critical binding site, and IL-8 binding affinity to the receptor N-domain is in the same concentration range as the IL-8 monomerdimer equilibrium. We observed that only the IL-8 monomer, and not the dimer, is competent in binding the receptor N-domain. Based on our results, we propose that IL-8 dimerization functions as a negative regulator for the receptor function and as a positive regulator for binding to glycosaminoglycans and that both play a role in the neutrophil recruitment process.
Journal of Biological Chemistry 09/2004; 279(35):36175-8. · 4.77 Impact Factor
