Abstract/Details

In-cell and in vitro studies of disease related protein-protein interactions using NMR-spectroscopy


2011 2011

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Abstract (summary)

The receptor for advanced glycation end products (RAGE) is a multiligand cell surface macromolecule that plays a central role in the etiology of diabetes, inflammation, and neurodegeneration. The cytoplasmic domain of RAGE, ctRAGE, is critical for RAGE-dependent signal transduction. As the most membrane proximal event, mDia1 binds to ctRAGE and is essential for RAGE ligand-stimulated phosphorylation of AKT and cell proliferation/migration. We show that ctRAGE contains an unusual alpha-turn that mediates the mDia1-ctRAGE interaction and is required for RAGE dependent signaling. The results establish a novel mechanism through which an extracellular signal initiated by RAGE ligands regulates RAGE signaling in a manner requiring mDia1.

Interactions between biological macromolecules give rise to and regulate biological activity. This activity is manifest through structural dynamics and changes in the macromolecular structures that comprise these interactions. Until recently, mostly in vitro techniques have been used to study macromolecular interactions that govern biological processes under conditions remote from those existing in the cell. With the advent of in-cell Nuclear Magnetic Resonance (NMR) spectroscopy, these processes can now be studied within a cellular environment. Here we review the latest techniques in order to study proteins under more physiological relevant conditions.

Mycobacterium tuberculosis ability to resist nitric oxide stress, from the immune initiated response, relies on then bacteria's ability to target misfolded or damaged proteins. Mycobacterium tuberculosis uses (Prokaryotic Ubiquitin like protein) Pup to target proteins for proteasome mediated degradation. Proteins which have been tagged by Pup interact with mycobacterial ATPase (Mpa). Using STINT NMR we characterized the interactions between Pup and Mpa in-cell and show that Pup's C- terminus is engaged by Mpa. Pups N terminus was also shown to be engaged only in the presence of the Mpa-proteasome complex. We also demonstrate, through various biochemical assays that Pup which is not deamidated is not a substrate for proteasomal degradation.

Indexing (details)


Subject
Analytical chemistry;
Biochemistry
Classification
0486: Analytical chemistry
0487: Biochemistry
Identifier / keyword
Pure sciences; Protein-protein interactions; Receptor for advanced glycation end products; Ubiquitin
Title
In-cell and in vitro studies of disease related protein-protein interactions using NMR-spectroscopy
Author
Maldonado, Andres Y.
Number of pages
166
Publication year
2011
Degree date
2011
School code
0668
Source
DAI-B 73/04, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
ISBN
9781267104175
Advisor
Shekhtman, Alexander
Committee member
Belfort, Marlene; Pande, Jayanti; Scholes, Charles P.; Shekhtman, Alexander; Theimer, Carla
University/institution
State University of New York at Albany
Department
Chemistry
University location
United States -- New York
Degree
Ph.D.
Source type
Dissertations & Theses
Language
English
Document type
Dissertation/Thesis
Dissertation/thesis number
3490106
ProQuest document ID
916764113
Copyright
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
http://search.proquest.com/docview/916764113
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