Characterization of Mutants of Human Small Heat Shock Protein HspB1 Carrying Replacements in the N-Terminal Domain and Associated with Hereditary Motor Neuron Diseases

PLoS One. 2015 May 12;10(5):e0126248. doi: 10.1371/journal.pone.0126248. eCollection 2015.

Abstract

Physico-chemical properties of the mutations G34R, P39L and E41K in the N-terminal domain of human heat shock protein B1 (HspB1), which have been associated with hereditary motor neuron neuropathy, were analyzed. Heat-induced aggregation of all mutants started at lower temperatures than for the wild type protein. All mutations decreased susceptibility of the N- and C-terminal parts of HspB1 to chymotrypsinolysis. All mutants formed stable homooligomers with a slightly larger apparent molecular weight compared to the wild type protein. All mutations analyzed decreased or completely prevented phosphorylation-induced dissociation of HspB1 oligomers. When mixed with HspB6 and heated, all mutants yielded heterooligomers with apparent molecular weights close to ~400 kDa. Finally, the three HspB1 mutants possessed lower chaperone-like activity towards model substrates (lysozyme, malate dehydrogenase and insulin) compared to the wild type protein, conversely the environmental probe bis-ANS yielded higher fluorescence with the mutants than with the wild type protein. Thus, in vitro the analyzed N-terminal mutations increase stability of large HspB1 homooligomers, prevent their phosphorylation-dependent dissociation, modulate their interaction with HspB6 and decrease their chaperoning capacity, preventing normal functioning of HspB1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • HSP20 Heat-Shock Proteins / chemistry
  • HSP20 Heat-Shock Proteins / genetics*
  • HSP20 Heat-Shock Proteins / metabolism
  • HSP27 Heat-Shock Proteins / chemistry
  • HSP27 Heat-Shock Proteins / genetics*
  • HSP27 Heat-Shock Proteins / metabolism
  • Heat-Shock Proteins
  • Humans
  • Molecular Chaperones / chemistry
  • Molecular Chaperones / genetics
  • Motor Neurons / metabolism*
  • Motor Neurons / pathology
  • Muscular Atrophy, Spinal / drug therapy
  • Muscular Atrophy, Spinal / genetics*
  • Muscular Atrophy, Spinal / pathology
  • Mutation
  • Phosphorylation
  • Protein Aggregation, Pathological / genetics
  • Protein Binding / genetics
  • Protein Multimerization / genetics
  • Protein Structure, Quaternary
  • Proteolysis

Substances

  • HSP20 Heat-Shock Proteins
  • HSP27 Heat-Shock Proteins
  • HSPB1 protein, human
  • HSPB6 protein, human
  • Heat-Shock Proteins
  • Molecular Chaperones

Grants and funding

Investigation of physico-chemical properties of isolated mutants performed by LKM and NBG was supported by Russian Science Foundation (RSF) (grant No 14-35-00026). Protein-protein interaction and bioinformatic analysis performed by SDW and SVS were supported by Flanders Research Foundation (grant G.06978.08 to SVS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.