Phosphorylation dependence of hsp27 multimeric size and molecular chaperone function.

Area of Science:

• Biochemistry
• Molecular Biology
• Protein Folding

Background:

• Heat shock protein 27 (Hsp27) functions as a molecular chaperone, existing as large oligomers.
• Phosphorylation at specific serine residues (Ser-15, -78, -82) is known to disassemble Hsp27 oligomers.
• The relative in vitro chaperone activity of unphosphorylated Hsp27 versus its triple Ser-to-Asp phospho-mimic mutant remains controversial.

Purpose of the Study:

• To investigate the relationship between Hsp27 self-association and its chaperone activity.
• To compare the chaperone function and self-association of phosphorylated Hsp27 with its phospho-mimic mutant.
• To determine the most active form of Hsp27 for preventing protein aggregation.

Main Methods:

• Analytical ultracentrifugation was used to monitor Hsp27 self-association.
• Chaperone activity was assessed by measuring the aggregation of reduced insulin and alpha-lactalbumin.
• Hsp27 was phosphorylated using MAPKAP-2 kinase, and phosphorylation levels were tracked by urea-PAGE.

Main Results:

• A decrease in Hsp27 self-association correlated with increased suppression of insulin and alpha-lactalbumin aggregation.
• Phosphorylated Hsp27 exhibited the least self-association and highest chaperone activity, followed by the phospho-mimic, then unphosphorylated Hsp27.
• Hsp27 inhibited the formation of larger aggregates from pre-aggregated insulin, independent of phosphorylation status.

Conclusions:

• The Hsp27 dimer represents the most active molecular chaperone for insulin and alpha-lactalbumin under physiological conditions.
• The Hsp27 phospho-mimic mutant only partially replicates the self-association and chaperone function of phosphorylated Hsp27.
• Two models were proposed to explain the chaperone activity of Hsp27 dimer in inhibiting insulin aggregation.