Structural and functional consequences of NEDD8 phosphorylation

Nat Commun. 2021 Oct 12;12(1):5939. doi: 10.1038/s41467-021-26189-9.

Abstract

Ubiquitin (Ub) and Ub-like proteins (Ubls) such as NEDD8 are best known for their function as covalent modifiers of other proteins but they are also themselves subject to post-translational modifications including phosphorylation. While functions of phosphorylated Ub (pUb) have been characterized, the consequences of Ubl phosphorylation remain unclear. Here we report that NEDD8 can be phosphorylated at S65 - the same site as Ub - and that S65 phosphorylation affects the structural dynamics of NEDD8 and Ub in a similar manner. While both pUb and phosphorylated NEDD8 (pNEDD8) can allosterically activate the Ub ligase Parkin, they have different protein interactomes that in turn are distinct from those of unmodified Ub and NEDD8. Among the preferential pNEDD8 interactors are HSP70 family members and we show that pNEDD8 stimulates HSP70 ATPase activity more pronouncedly than unmodified NEDD8. Our findings highlight the general importance of Ub/NEDD8 phosphorylation and support the notion that the function of pUb/pNEDD8 does not require their covalent attachment to other proteins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • Amino Acid Sequence
  • Cloning, Molecular
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • HEK293 Cells
  • HSP70 Heat-Shock Proteins / chemistry
  • HSP70 Heat-Shock Proteins / classification
  • HSP70 Heat-Shock Proteins / genetics
  • HSP70 Heat-Shock Proteins / metabolism*
  • Humans
  • Kinetics
  • NEDD8 Protein / chemistry
  • NEDD8 Protein / genetics
  • NEDD8 Protein / metabolism*
  • Phosphorylation
  • Protein Binding
  • Protein Interaction Mapping
  • Protein Processing, Post-Translational*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Substrate Specificity
  • Ubiquitin / chemistry
  • Ubiquitin / genetics
  • Ubiquitin / metabolism*
  • Ubiquitination

Substances

  • HSP70 Heat-Shock Proteins
  • NEDD8 Protein
  • NEDD8 protein, human
  • Recombinant Proteins
  • Ubiquitin