Intrinsic Disorder to Order Transitions in the Scaffold Phosphoprotein P from the Respiratory Syncytial Virus RNA Polymerase Complex

Biochemistry. 2016 Mar 15;55(10):1441-54. doi: 10.1021/acs.biochem.5b01332. Epub 2016 Mar 4.

Abstract

Intrinsic disorder is at the center of biochemical regulation and is particularly overrepresented among the often multifunctional viral proteins. Replication and transcription of the respiratory syncytial virus (RSV) relies on a RNA polymerase complex with a phosphoprotein cofactor P as the structural scaffold, which consists of a four-helix bundle tetramerization domain flanked by two domains predicted to be intrinsically disordered. Because intrinsic disorder cannot be reduced to a defined atomic structure, we tackled the experimental dissection of the disorder-order transitions of P by a domain fragmentation approach. P remains as a tetramer above 70 °C but shows a pronounced reversible secondary structure transition between 10 and 60 °C. While the N-terminal module behaves as a random coil-like IDP in a manner independent of tetramerization, the isolated C-terminal module displays a cooperative and reversible metastable transition. When linked to the tetramerization domain, the C-terminal module becomes markedly more structured and stable, with strong ANS binding. Therefore, the tertiary structure in the C-terminal module is not compact, conferring "late" molten globule-like IDP properties, stabilized by interactions favored by tetramerization. The presence of a folded structure highly sensitive to temperature, reversibly and almost instantly formed and broken, suggests a temperature sensing activity. The marginal stability allows for exposure of protein binding sites, offering a thermodynamic and kinetic fine-tuning in order-disorder transitions, essential for the assembly and function of the RSV RNA polymerase complex.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA-Directed RNA Polymerases / chemistry
  • DNA-Directed RNA Polymerases / metabolism*
  • Humans
  • Nuclear Matrix-Associated Proteins / chemistry
  • Nuclear Matrix-Associated Proteins / metabolism*
  • Phosphoproteins / chemistry
  • Phosphoproteins / metabolism*
  • Protein Binding / physiology
  • Respiratory Syncytial Virus, Human / chemistry
  • Respiratory Syncytial Virus, Human / metabolism*
  • Viral Proteins / chemistry
  • Viral Proteins / metabolism

Substances

  • Nuclear Matrix-Associated Proteins
  • Phosphoproteins
  • Viral Proteins
  • DNA-Directed RNA Polymerases