Expression and isotopic labeling of structural domains of the human protein DEK

Matthew Devany, N. Prasad Kotharu, Hiroshi Matsuo

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

The 375 amino acid human protein DEK has been expressed in two functional, structured domains. DEK is an abundant nuclear protein that associates with chromatin and alters its topology by introducing positive supercoiling in DNA, which results in lower replication efficiency. DEK has clinical importance as transfection of the cDNA of the C-terminal region of DEK can partially reverse the abnormal DNA-mutagen sensitivity in fibroblasts derived from ataxia-telangiectasia (A-T) patients, and elevated levels of DEK mRNA are observed in various forms of cancer. Because high-level expression of full-length DEK has proved elusive, we sought an alternative for structural studies that would provide insights on DEK's function. We have discovered that DEK contains two structured domains and have expressed these domains at a high level in Escherichia coli in M9 minimal media. The N-terminal domain (amino acids 68-226) includes the region responsible for introducing supercoils into DNA, and the C-terminal domain (amino acids 309-375) includes the region that can reverse the abnormal DNA-mutagen sensitivity of A-T cells. 1H-15N correlation nuclear magnetic resonance spectra of these two fragments reveal the characteristic signature of folded proteins.

Original languageEnglish (US)
Pages (from-to)244-247
Number of pages4
JournalProtein Expression and Purification
Volume40
Issue number2
DOIs
StatePublished - Apr 2005

Bibliographical note

Funding Information:
NMR instrumentation was provided with funds from the NSF (BIR-961477), the University of Minnesota Medical School, and the Minnesota Medical Foundation. We thank Basic Science Computer Laboratory of University of Minnesota for providing us SGI workstations. This work is partially supported by a grant from Grant-in-Aid of Research, Artistry, and Scholarship from the Graduate School of University of Minnesota to H.M. M.D. was partially supported by the National Institute of Health Predoctoral Training Grant.

Keywords

  • Acute myelogenous leukemia
  • Ataxiatelangiectasia
  • DEK
  • Proto-oncogene protein

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