含聚腺苷酸核糖核蛋白在寡聚胸苷酸纤维素上的亲和层析
Affinity chromatography of poly(adenylic acid)-containing ribonucleoproteins on oligothymidylic acid-cellulose.
作者信息
De Meyer P, De Herdt E, Kondo M, Slegers H
出版信息
J Biochem Biophys Methods. 1980 May;2(5):311-4. doi: 10.1016/0165-022x(80)90056-1.
Binding of poly(A)-containing RNP to oligo(dT)-cellulose has been investigated as a function of mono- and divalent ion concentration. 80-90% binding was obtained either in high (500 mM) or in moderate NaCl concentrations in the presence of 5 mM MgCl2. At 40 mM NaCl and 5 mM MgCl2 poly(A)+-RNP exhibit approximately the same stability as poly(A)+-RNA in binding to oligo(dT)-cellulose with a melting temperature of 41 and 45 degrees C, respectively, indicating that the protein moiety has no effect on the ribonucleoprotein binding in these conditions. Differences were observed in the elution of poly(A)+-RNA and poly(A)+-RNP from oligo(dT)-cellulose in buffer without salts. Poly(A)+-RNA was completely removed at 4 degrees C whereas the melting temperature of poly(A)+-RNP was only decreased to 34 degrees C. The isolation of poly(A)+-RNP by thermal elution from oligo(dT)-cellulose is described.
已研究了含聚腺苷酸（poly(A)）的核糖核蛋白（RNP）与寡聚（dT）纤维素的结合情况，该结合是单价和二价离子浓度的函数。在5 mM氯化镁存在的情况下，无论是在高浓度（500 mM）还是中等浓度的氯化钠中，均可实现80 - 90%的结合。在40 mM氯化钠和5 mM氯化镁条件下，聚腺苷酸阳性（poly(A)+）-RNP与聚腺苷酸阳性（poly(A)+）-RNA在结合寡聚（dT）纤维素时表现出大致相同的稳定性，其解链温度分别为41摄氏度和45摄氏度，这表明在这些条件下蛋白质部分对核糖核蛋白的结合没有影响。在无盐缓冲液中，观察到聚腺苷酸阳性（poly(A)+）-RNA和聚腺苷酸阳性（poly(A)+）-RNP从寡聚（dT）纤维素上洗脱时存在差异。聚腺苷酸阳性（poly(A)+）-RNA在4摄氏度时被完全洗脱，而聚腺苷酸阳性（poly(A)+）-RNP的解链温度仅降至34摄氏度。本文描述了通过热洗脱从寡聚（dT）纤维素中分离聚腺苷酸阳性（poly(A)+）-RNP的方法。
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