Mimicking phosphorylation of alphaB-crystallin affects its chaperone activity.
AlphaB-crystallin is a member of the sHsp (small heat-shock protein) family that prevents misfolded target proteins from aggregating and precipitating. Phosphorylation at three serine residues (Ser19, Ser45 and Ser59) is a major post-translational modification that occurs to alphaB-crystallin. In the present study, we produced recombinant proteins designed to mimic phosphorylation of alphaB-crystallin by incorporating a negative charge at these sites. We employed these mimics to undertake a mechanistic and structural investigation of the effect of phosphorylation on the chaperone activity of alphaB-crystallin to protect against two types of protein misfolding, i.e. amorphous aggregation and amyloid fibril assembly. We show that mimicking phosphorylation of alphaB-crystallin results in more efficient chaperone activity against both heat-induced and reduction-induced amorphous aggregation of target proteins. Mimick-ing phosphorylation increased the chaperone activity of alphaB-crystallin against one amyloid-forming target protein (kappa-casein), but decreased it against another (ccbeta-Trp peptide). We observed that both target protein identity and solution (buffer) conditions are critical factors in determining the relative chaperone ability of wild-type and phosphorylated alphaB-crystallins. The present study provides evidence for the regulation of the chaperone activity of alphaB-crystallin by phosphorylation and indicates that this may play an important role in alleviating the pathogenic effects associated with protein conformational diseases.
Ecroyd
Heath
H
School of Chemistry and Physics, University of Adelaide, Adelaide, SA 5005, Australia.
Meehan
Sarah
S
Horwitz
Joseph
J
Aquilina
J Andrew
JA
Benesch
Justin L P
JL
Robinson
Carol V
CV
Macphee
Cait E
CE
Carver
John A
JA
D000818:Animals; D002364:Caseins / Q000302:isolation & purification; D002374:Catalase / Q000378:metabolism; D002417:Cattle; D005260:Female; D007768:Lactalbumin / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D008892:Milk; D018832:Molecular Chaperones / Q000502:physiology; D009391:Nephelometry and Turbidimetry; D010766:Phosphorylation; D012542:Scattering, Radiation; D038203:alpha-Crystallin B Chain / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D002364:Caseins; D018832:Molecular Chaperones; D038203:alpha-Crystallin B Chain; D007768:Lactalbumin; D002374:Catalase