Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky pr. 33, Moscow 119071, Russia.
Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky pr. 33, Moscow 119071, Russia.
Int J Biol Macromol. 2015 May;76:86-93. doi: 10.1016/j.ijbiomac.2015.02.022. Epub 2015 Feb 20.
The main function of small heat shock proteins acting as suppressors of aggregation of non-native proteins is greatly influenced by crowded environment in the cell and the presence of divalent metal ions. The goal of the present work was to study the effects of Ca(2+) and Mg(2+) ions on the quaternary structure and anti-aggregation activity of αB-crystallin under crowding conditions. We showed that Ca(2+) and Mg(2+) ions induced formation of suboligomeric forms of αB-crystallin. This effect was retained in the presence of crowder (polyethylene glycol), although to a lesser degree. The chaperone-like activity of αB-crystallin was analyzed using heat-induced aggregation of myosin subfragment 1 (S1) at 40°C. In the absence of crowding agents chaperone-like activity of αB-crystallin exhibited low sensitivity to the presence of Ca(2+) and Mg(2+) ions. The addition of the crowding agents (polyethylene glycol 20000, Ficoll 70) dramatically increased S1 aggregation rates and significantly depressed anti-aggregation activity of αB-crystallin. Low concentrations of Ca(2+) (0.1mM) and Mg(2+) (10mM) partially restored the chaperone-like activity of αB-crystallin in the presence of crowders. This effect was observed at relatively low values of [αB-crystallin]/[S1] molar ratio, however, at [αB-crystallin]/[S1]>0.2 the stimulating effect of Ca(2+) became less pronounced. These findings might indicate that under crowded cell conditions different factors, including divalent cations, can effectively modulate chaperone-like activity of protein chaperones, which otherwise cannot properly cope with crowding-provoked accelerated rates of substrates aggregation.
小分子热休克蛋白作为非天然蛋白质聚集抑制剂的主要功能受到细胞拥挤环境和二价金属离子存在的极大影响。本工作的目的是研究 Ca(2+)和 Mg(2+)离子对拥挤条件下 αB-晶体蛋白的四级结构和抗聚集活性的影响。我们表明，Ca(2+)和 Mg(2+)离子诱导 αB-晶体蛋白形成亚寡聚形式。这种效应在存在拥挤剂（聚乙二醇）的情况下仍然存在，尽管程度较小。使用 40°C 下肌球蛋白小片段 1 (S1)的热诱导聚集来分析 αB-晶体蛋白的伴侣样活性。在不存在拥挤剂的情况下，αB-晶体蛋白的伴侣样活性对 Ca(2+)和 Mg(2+)离子的存在表现出低敏感性。添加拥挤剂（聚乙二醇 20000、Ficoll 70）会显著增加 S1 聚集速率，并显著降低 αB-晶体蛋白的抗聚集活性。低浓度的 Ca(2+)（0.1mM）和 Mg(2+)（10mM）在拥挤剂存在下部分恢复了 αB-晶体蛋白的伴侣样活性。然而，在[αB-晶体蛋白]/[S1]摩尔比相对较低的情况下，观察到这种效应，但是在[αB-晶体蛋白]/[S1]>0.2 时，Ca(2+)的刺激作用变得不那么明显。这些发现可能表明，在拥挤的细胞条件下，包括二价阳离子在内的不同因素可以有效地调节蛋白质伴侣的伴侣样活性，否则它们无法很好地应对拥挤引起的底物聚集加速速率。
