del Valle Luis J, Escribano Cristina, Pérez Juan J, Garriga Pere
Departament d'Enginyeria Química, Universitat Politècnica de Catalunya, Colom 1, 08222 Terrassa, Catalonia, Spain.
Biochim Biophys Acta. 2002 Nov 19;1601(1):100-9. doi: 10.1016/s1570-9639(02)00429-6.
Alpha-crystallin, one of the major proteins in the vertebrate eye lens, acts as a molecular chaperone, like the small heat-shock proteins, by protecting other proteins from denaturing under stress or high temperature conditions. alpha-Crystallin aggregation is involved in lens opacification, and high [Ca(2+)] has been associated with cataract formation, suggesting a role for this cation in the pathological process. We have investigated the effect of Ca(2+) on the thermal stability of alpha-crystallin by UV and Fourier-transform infrared (FTIR) spectroscopies. In both cases, a Ca(2+)-induced decrease in the midpoint of the thermal transition is detected. The presence of high [Ca(2+)] results also in a marked decrease of its chaperone activity in an insulin-aggregation assay. Furthermore, high Ca(2+) concentration decreases Cys reactivity towards a sulfhydryl reagent. The results obtained from the spectroscopic analysis, and confirmed by circular dichroism (CD) measurements, indicate that Ca(2+) decreases both secondary and tertiary-quaternary structure stability of alpha-crystallin. This process is accompanied by partial unfolding of the protein and a clear decrease in its chaperone activity. It is concluded that Ca(2+) alters the structural stability of alpha-crystallin, resulting in impaired chaperone function and a lower protective ability towards other lens proteins. Thus, alpha-crystallin aggregation facilitated by Ca(2+) would play a role in the progressive loss of transparency of the eye lens in the cataractogenic process.
α-晶状体蛋白是脊椎动物眼晶状体中的主要蛋白质之一，它作为分子伴侣，类似于小分子热休克蛋白，通过保护其他蛋白质在应激或高温条件下不发生变性。α-晶状体蛋白的聚集与晶状体混浊有关，高[Ca(2+)]与白内障形成相关，提示该阳离子在病理过程中起作用。我们通过紫外和傅里叶变换红外（FTIR）光谱研究了Ca(2+)对α-晶状体蛋白热稳定性的影响。在这两种情况下，均检测到Ca(2+)诱导的热转变中点降低。在胰岛素聚集试验中，高[Ca(2+)]的存在也导致其伴侣活性显著降低。此外，高Ca(2+)浓度降低了半胱氨酸对巯基试剂的反应性。光谱分析结果并经圆二色性（CD）测量证实，表明Ca(2+)降低了α-晶状体蛋白的二级和三级-四级结构稳定性。这一过程伴随着蛋白质的部分解折叠及其伴侣活性的明显降低。结论是Ca(2+)改变了α-晶状体蛋白的结构稳定性，导致伴侣功能受损以及对其他晶状体蛋白的保护能力降低。因此，Ca(2+)促进的α-晶状体蛋白聚集在白内障形成过程中晶状体透明度的逐渐丧失中起作用。
