Ma Z, Hanson S R, Lampi K J, David L L, Smith D L, Smith J B
Department of Chemistry, University of Nebraska, Lincoln 68588, USA.
Exp Eye Res. 1998 Jul;67(1):21-30. doi: 10.1006/exer.1998.0482.
Analysis of water-soluble crystallins from human lenses, ages 32 week fetal to 55 years has led to identification of the major modifications of the proteins comprising the lens. These modifications were identified by the masses of the proteins determined by electrospray ionization mass spectrometry after the proteins were separated by gel filtration and reversed phase high performance liquid chromatography. Examination of all the proteins isolated from the water soluble portion demonstrated that the major age-related modifications causing significant alteration in the molecular weights of the lens crystallins include truncation of the N-termini of beta B1, beta A3 and beta A1, and partial phosphorylation and C-terminal degradation of alpha-crystallins. N-terminal degradation of beta B1, beta A3 and beta A1 was evident in human lenses less than one year old, and the proportion of these truncated proteins became greater with age. Phosphorylation of alpha A- and alpha B-crystallins increased from the fetal to the 3 year old lens, but did not change with further aging. Minor components indicating truncation of the C-termini of alpha-crystallins were found in older lenses. In contrast to beta B1, beta A3 and beta A1, the masses of the major species of alpha A, alpha B, beta B2, beta A4, gamma S, gamma C, and gamma D did not change with aging. This suggested that the major modifications to these crystallins are limited to deamidation and possibly intra-molecular disulfide bonds. These data, in conjunction with the data in the accompanying manuscript, established deamidation as a common modification, since deamidation, which causes only a one dalton change in mass, is the only modification that is consistent with the absence of a detectable change in molecular weight and the observed increased acidity demonstrated in the two-dimensional gels of the accompanying paper. Other age related changes included a decrease in beta B3 (M(r) 24224), a major component of the fetal lens, which was not detected in lenses older than 3 years, and increases in the ratios of alpha B:alpha A and gamma S:gamma C.
对年龄从32周胎儿到55岁的人晶状体水溶性晶状体蛋白进行分析，已确定了构成晶状体的蛋白质的主要修饰。这些修饰是通过凝胶过滤和反相高效液相色谱分离蛋白质后，用电喷雾电离质谱法测定蛋白质质量来确定的。对从水溶性部分分离出的所有蛋白质进行检查表明，导致晶状体蛋白分子量发生显著变化的主要与年龄相关的修饰包括βB1、βA3和βA1的N端截短，以及α晶状体蛋白的部分磷酸化和C端降解。βB1、βA3和βA1的N端降解在1岁以下的人晶状体中很明显，并且这些截短蛋白的比例随年龄增长而增加。αA-和αB-晶状体蛋白的磷酸化从胎儿期到3岁晶状体增加，但随着年龄进一步增长没有变化。在老年晶状体中发现了表明α晶状体蛋白C端截短的次要成分。与βB1、βA3和βA1相反，αA、αB、βB2、βA4、γS、γC和γD的主要种类的质量不随年龄变化。这表明这些晶状体蛋白的主要修饰仅限于脱酰胺作用以及可能的分子内二硫键。这些数据与随附手稿中的数据一起，将脱酰胺作用确立为一种常见修饰，因为脱酰胺作用仅导致质量上1道尔顿的变化，是唯一与分子量无 detectable 变化以及随附论文二维凝胶中观察到的酸度增加相一致的修饰。其他与年龄相关的变化包括胎儿晶状体的主要成分βB3（M(r) 24224）减少，在3岁以上的晶状体中未检测到，以及αB:αA和γS:γC的比例增加。
