Cloning and Biochemical Analysis of the Ataxia-Telangiectasia (A-T) Gene in Xenopus Laevis
Abstract
The ATM protein, encoded by the gene responsible for the human genetic disorder ataxia telangiectasia (A-T), is a 370 KDa protein with a COOH-terminal domain similar to the catalytic subunit of the phosphoinositide 3 kinases (PI 3-kinases). This places ATM within a family of protein kinases that function in maintenance of genome stability, in cell cycle control, and cellular responses to DNA damage. In order to address its proposed function in cell cycle regulation and DNA damage checkpoints, we propose to use the xenopus system for analysis of these biochemical functions. We have cloned the xenopus homologue of the human ATM gene and are in the process of analyzing its biochemical function. To this end we have successfully produced recombinant fragments of the ATM protein, including the kinase domain, in E. coli and by in vitro translation in reticulocyte lysates. In addition, we have raised polyclonal antibodies against recombinant protein made in E. coli. The antibody recognizes a single band of approximately 350 KDa on western blots from extracts of xenopus eggs and embryos.
Document Details
- Document Type
- Technical Report
- Publication Date
- Sep 01, 1998
- Accession Number
- ADA365457
Entities
People
- Carmel Hensey
Organizations
- Columbia University