Synthesis and characterization of a peptide identified as a functional element in αA-crystallin

被引:196
作者
Sharma, KK [1 ]
Kumar, RS
Kumar, GS
Quinn, PT
机构
[1] Univ Missouri, Dept Ophthalmol, Mason Eye Inst, Columbia, MO 65212 USA
[2] Univ Missouri, Dept Biochem, Columbia, MO 65212 USA
关键词
D O I
10.1074/jbc.275.6.3767
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Eye lens alpha-crystallin is a member of the small heat shock protein (sHSP) family and forms large multimeric structures. Earlier studies have shown that it can act like a molecular chaperone and form a stable complex with partially unfolded proteins, We have observed that prior binding of the hydrophobic protein melittin to alpha-crystallin diminishes its chaperone-like activity toward denaturing alcohol dehydrogenase, suggesting the presence of mutually exclusive sites for these proteins in alpha-crystallin, To investigate the mechanism of the interaction between alpha-crystallin and substrate proteins, we determined the melittin-binding sites in alpha-crystallin by cross-linking studies. Localization of melittin-binding sites in alpha-crystallin resulted in the identification of RTLGPFYPSR and FVIFLDVKHFSPEDLTVK of alpha A-crystallin and FSVNLDVK of alpha B-crystallin as the chaperone sites. Of these sites, FVIFLDVKHFSPEDLTVK and FSVNLDVK were identified earlier as 1,1'-bi(4-anilino) naphthalene-5,5'-disulfonic acid (bis-ANS)-binding hydrophobic sites. Here we also report the synthesis and characterization of the peptide, KFVIFLDVKHFSPEDLTVK, having the melittin as well as bis-ANS-binding sequence of alpha A-crystallin. We show that this peptide has characteristics similar to that of alpha A-crystallin by in vitro thermal aggregation assay, gel filtration study, CD spectroscopy, and bis-ANS interaction studies. The peptide sequence corresponds to the beta 3 and beta 4 region present in the cu-crystallin domain of sHSP 16.5. We hypothesize that the alpha-crystallin domain in other sHSPs may have a similar function and would likely possess the anti-aggregation property even when separated from the native protein.
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收藏
页码:3767 / 3771
页数:5
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