The mutation Asp69-->Ser affects the chaperone-like activity of alpha A-crystallin.
alpha-Crystallins are members of the family of small heat-shock proteins. The conformation and mode of action of these 'junior chaperones' are unknown. To investigate the structure and chaperone-like activity, four mutants of bovine alpha A-crystallin were generated by site-directed mutagenesis. In comparison with wild-type alpha A-crystallin, the D69S mutant, in which a highly conserved charged residue has been replaced, forms larger multimers and displays a threefold reduced heat-protection capacity. The conformation and thermal stability of this mutant are not noticeably affected. Three other mutations, replacing hydrophobic by uncharged hydrophilic residues, were aimed at disturbing hydrophobic intersubunit interactions. None of these mutations resulted in major structural perturbations and only minor differences in heat-protective capacity were observed. Although it is assumed that small heat-shock proteins interact with denaturing proteins via their hydrophobic surfaces, this study clearly shows that charged residues in alpha-crystallin can also influence the efficiency of substrate binding.
Smulders
R H
RH
Department of Biochemistry, University of Nijmegen, The Netherlands.
Merck
K B
KB
Aendekerk
J
J
Horwitz
J
J
Takemoto
L
L
Slingsby
C
C
Bloemendal
H
H
De Jong
W W
WW
D000595:Amino Acid Sequence; D001216:Asparagine / Q000235:genetics*; D001483:Base Sequence; D002850:Chromatography, Gel / Q000379:methods; D003459:Crystallins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics; D005453:Fluorescence; D006358:Hot Temperature; D018832:Molecular Chaperones / Q000502:physiology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011487:Protein Conformation; D017510:Protein Folding; D012694:Serine / Q000235:genetics*; D013329:Structure-Activity Relationship; D013499:Surface Properties
D003459:Crystallins; D018832:Molecular Chaperones; D012694:Serine; D001216:Asparagine