Association of RNA with the uracil-DNA-degrading factor has major conformational effects and is potentially involved in protein folding

Angela Bekesi, Maria Pukancsik, Peter Haasz, Lilla Felfoldi, Ibolya Leveles, Villo Muha, Eva Hunyadi-Gulyas, Anna Erdei, Katalin F. Medzihradszky, Beata G. Vertessy

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Recently, a novel uracil-DNA-degrading factor protein (UDE) was identified in Drosophila melanogaster, with homologues only in pupating insects. Its unique uracil-DNA-degrading activity and a potential domain organization pattern have been described. UDE seems to be the first representative of a new protein family with unique enzyme activity that has a putative role in insect development. In addition, UDE may also serve as potential tool in molecular biological applications. Owing to lack of homology with other proteins with known structure and/or function, de novo data are required for a detailed characterization of UDE structure and function. Here, experimental evidence is provided that recombinant protein is present in two distinct conformers. One of these contains a significant amount of RNA strongly bound to the protein, influencing its conformation. Detailed biophysical characterization of the two distinct conformational states (termed UDE and RNA-UDE) revealed essential differences. UDE cannot be converted into RNA-UDE by addition of the same RNA, implying putatively joint processes of RNA binding and protein folding in this conformational species. By real-time PCR and sequencing after random cloning, the bound RNA pool was shown to consist of UDE mRNA and the two ribosomal RNAs, also suggesting cotranslational RNA-assisted folding. This finding, on the one hand, might open a way to obtain a conformationally homogeneous UDE preparation, promoting successful crystallization; on the other hand, it might imply a further molecular function of the protein. In fact, RNA-dependent complexation of UDE was also demonstrated in a fruit fly pupal extract, suggesting physiological relevance of RNA binding of this DNA-processing enzyme.

Original languageEnglish
Pages (from-to)295-315
Number of pages21
JournalFEBS Journal
Volume278
Issue number2
DOIs
Publication statusPublished - Jan 1 2011

    Fingerprint

Keywords

  • RNA binding
  • RNA-assisted folding
  • conformational states
  • cotranslational folding
  • uracil-DNA-degrading factor

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this