Deletion of 54FLRAPSWF61 Residues Decreases the Oligomeric Size and Enhances the Chaperone Function of αB-Crystallin

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• 作者：Puttur Santhoshkumar ; Raju Murugesan ; K. Krishna Sharma
• 刊名：Biochemistry
• 出版年：2009
• 出版时间：June 16, 2009
• 年：2009
• 卷：48
• 期：23
• 页码：5066-5073
• 全文大小：882K
• 年卷期：v.48,no.23(June 16, 2009)
• ISSN：1520-4995

文摘

αB-Crystallin is a member of the small heat shock protein family and is known to have chaperone activity. Using a peptide scan approach, we previously determined that regions 42−57, 60−71, and 88−123 in αB-crystallin interact with αA-crystallin during heterooligomer formation. To further characterize the significance of the N-terminal domain of αB-crystallin, we prepared a deletion mutant that lacks residues ⁵⁴FLRAPSWF⁶¹ (αBΔ54−61) and found that the absence of residues 54−61 in αB-crystallin significantly decreased the homooligomeric mass of αB-crystallin. The average oligomeric mass of wild-type αB-crystallin and of αBΔ54−61, calculated using multiangle light scattering, was 624 and 382 kDa, respectively. The mutant subunits aggregate to form smaller, less-compact oligomers with a 4-fold increase in subunit exchange rate. Deletion of the 54−61 region resulted in a 50% decrease in intrinsic tryptophan fluorescence. The αBΔ54−61 mutant showed a 2-fold increase in 1,1′-bi(4-anilino)naphthalene-5,5′-disulfonic acid (bis-ANS) binding as compared to the wild-type protein, suggesting increased hydrophobicity of the mutant protein. Accompanying the evidence of increased hydrophobicity in the deletion mutant was a 10-fold increase in antiaggregation activity. Homooligomers of 6HαA (750 kDa) readily exchanged subunits with αBΔ54−61 homooligomers at 37 °C, forming heterooligomers with an intermediate mass of 625 kDa. Our data suggest that residues ⁵⁴FLRAPSWF⁶¹ contribute to the higher order assembly of αB-crystallin oligomers. Residues ⁵⁴FLRAPSWF⁶¹ in αB-crystallin are not essential for target protein binding during chaperone action, but this region apparently has a role in the chaperone activity of native αB-crystallin.