Structural basis of nucleic-acid recognition and double-strand unwinding by the essential neuronal protein Pur-alpha

Elife. 2016 Jan 8;5:e11297. doi: 10.7554/eLife.11297.

Abstract

The neuronal DNA-/RNA-binding protein Pur-alpha is a transcription regulator and core factor for mRNA localization. Pur-alpha-deficient mice die after birth with pleiotropic neuronal defects. Here, we report the crystal structure of the DNA-/RNA-binding domain of Pur-alpha in complex with ssDNA. It reveals base-specific recognition and offers a molecular explanation for the effect of point mutations in the 5q31.3 microdeletion syndrome. Consistent with the crystal structure, biochemical and NMR data indicate that Pur-alpha binds DNA and RNA in the same way, suggesting binding modes for tri- and hexanucleotide-repeat RNAs in two neurodegenerative RNAopathies. Additionally, structure-based in vitro experiments resolved the molecular mechanism of Pur-alpha's unwindase activity. Complementing in vivo analyses in Drosophila demonstrated the importance of a highly conserved phenylalanine for Pur-alpha's unwinding and neuroprotective function. By uncovering the molecular mechanisms of nucleic-acid binding, this study contributes to understanding the cellular role of Pur-alpha and its implications in neurodegenerative diseases.

Keywords: 5q31.3 microdeletion syndrome; d. melanogaster; e. coli; ALS; DNA unwinding; DNA-/RNA-protein interaction; FXTAS; X-ray crystallography; biochemistry; biophysics; structural biology.

MeSH terms

  • Animals
  • Crystallography, X-Ray
  • DNA, Single-Stranded / chemistry*
  • DNA, Single-Stranded / metabolism*
  • Drosophila
  • Drosophila Proteins / chemistry*
  • Drosophila Proteins / metabolism*
  • Gene Deletion
  • Genetic Complementation Test
  • Models, Molecular
  • Nucleic Acid Conformation*
  • Protein Binding
  • Protein Conformation
  • Transcription Factors / chemistry*
  • Transcription Factors / metabolism*

Substances

  • DNA, Single-Stranded
  • Drosophila Proteins
  • Pur-alpha protein, Drosophila
  • Transcription Factors