Kisilevsky R, Weiler L, Treloar M
J Biol Chem. 1978 Oct 10;253(19):7101-8.
Optimal conditions for reductive alkylation of ribosomal proteins in their native and denatured states were examined. The relative accessibility of rat liver ribosomal proteins to reductive alkylation was then examined. Intact ribosomes were firs labeled with [14C]formaldehyde and NaBH4. The proteins were then separated from RNA, denatured in 6 M guanidine, and labeled again using formaldehyde and NaB3H4. The relative accessibility of individual proteins to labeling in the intact state could thus be determined from their 3H/14C ratios following separation by two-dimensional electrophoresis. The results suggest that proteins S6, S11, S26, L3, and L35 are less accessible to labeling while proteins S1, S15, L11, L12, L16, and L24 appear relatively more accessible. The accessibility of individual proteins in ribosomes in different conformational states were then compared. The results indicated that S3, L7, and L36 are likely to be involved in a structural difference when normal polysomes and normal monomers are compared. Also, that S26 and L35, and probably S3, S20, L7, L8, L24, L27, L28 and L34 appear to be involved in a ribosomal conformation change induced by ethionine intoxication.
研究了核糖体蛋白在天然状态和变性状态下进行还原烷基化的最佳条件。随后检测了大鼠肝脏核糖体蛋白对还原烷基化的相对可及性。完整核糖体首先用[14C]甲醛和硼氢化钠进行标记。然后将蛋白质与RNA分离，在6M胍中变性，再用甲醛和硼氢化三钠再次标记。通过二维电泳分离后，根据各蛋白质的3H/14C比值，即可确定完整状态下单个蛋白质对标记的相对可及性。结果表明，蛋白质S6、S11、S26、L3和L35较难被标记，而蛋白质S1、S15、L11、L12、L16和L24相对更容易被标记。随后比较了处于不同构象状态的核糖体中单个蛋白质的可及性。结果表明，当比较正常多核糖体和正常单体时，S3、L7和L36可能与结构差异有关。此外，S26和L35，可能还有S3、S20、L7、L8、L24、L27、L28和L34似乎与乙硫氨酸中毒诱导的核糖体构象变化有关。
