Construction of His-tagged ATR fusion proteins: working toward the identification of ATR-interacting proteins
Loading...
Authors
Sassack, Mike
Issue Date
2004
Type
Presentation
Language
en_US
Keywords
Alternative Title
Abstract
The protein kinase ATR is responsible for initiating signal
transmission at cell cycle checkpoints. ATR is a 303 kDa protein
containing a carboxy-terminus kinase domain related to PI-3 kinase,
and is homologous to the ATM gene product in human cells and the
rad3/MEC1 proteins in yeast. These proteins, together with DNA-PK,
are part of the family of PI-3 kinase-related kinases (PIKKs). All
members of this family play important roles in cell cycle checkpoints
and operate to permit cell survival following many forms of DNA
damage. Upon damage of DNA in eukaryotic cells, ATR undergoes a
dramatic intranuclear relocalization, translocating to nuclear foci that
represent sites of DNA damage and repair. The ultimate goals of this
research are to identify new checkpoint pathway substrates that ATR is
able to phosphorylate post-DNA damage, and to initiate the
identification of potential inhibitors of ATR function. This study
commenced by dividing ATR into seven fragments. These fragments
were subsequently cloned into a vector system and transformed in
order to generate ATR fusion proteins. The induction of these fusion
constructs, mediated by their screening and conditioning, is subject to a
wide variety of specifications. Because the optimal induction
conditions have yet to be discovered, fine-tuning of the experimental
techniques must continue in order to give this research a higher degree
of significance. However, once successfully induced, a series of cotransfection
experiments may be initiated, in which the activity of ATR,
plus several additional proteins can be monitored under condition of
DNA damage.
Description
1 broadside : ill.
Citation
Publisher
Kalamazoo College