Nuclease treatment results in high specific purification of Creutzfeldt-Jakob disease infectivity with a density characteristic of nucleic acid-protein complexes.
Nuclease treatment results in high specific purification of Creutzfeldt-Jakob disease infectivity with a density characteristic of nucleic acid-protein complexes.
复制标题
核酸酶处理可实现克雅氏病感染性的高度特异性纯化，并具有核酸-蛋白质复合物的密度特征。
DOI：
10.1007/bf01323166
复制
发表时间：
1990
影响因子：
2.7
通讯作者：
Manuelidis,L
中科院分区：
文献类型：
--
作者：
Sklaviadis,T;Akowitz,A;Manuelidis,EE;Manuelidis,L
部分纯化的克雅氏病（CJD）的代表性制剂，包括解聚的密度梯度级分，用各种核酸酶处理。核糖核酸酶以及用微球菌核酸酶进行的彻底的双酶联免疫吸附试验并没有显著降低感染性，但是导致了对核酸的7,000倍的感染性特异性纯化。受保护的核酸包括长度高达2，000个碱基的种类。核酸酶处理后，在蔗糖中以1.27 g/cm 3的密度与核酸-蛋白质复合物共迁移。在梯度步骤中也实现了实质性的特异性蛋白纯化（约11，000倍），其中70%的宿主Gp 34（“朊病毒蛋白”）以及其他游离蛋白与感染性分离。这些CJD纯化比先前在羊瘙痒症中获得的纯化更好，并且可能用于非宿主蛋白质和核酸种类的进一步研究。这些数据与CJD样药物由核酸-蛋白质复合物组成的假设一致。
Representative preparations of partially purified Creutzfeldt-Jakob disease (CJD), including disaggregated density gradient fractions, were treated with a variety of nucleases. RNases as well as exhaustive digestions with micrococcal nuclease did not significantly diminish infectivity, but resulted in an ∼ 7,000-fold specific purification of infectivity with respect of nucleic acid. Protected nucleic acids included species of up to 2,000 bases in length. After nuclease treatment, infectivity co-migrated with nucleic acid-protein complexes at a density of 1.27 g/cm3in sucrose. Substantial specific protein purification were also achieved in the gradient step (∼ 11,000-fold), where 70% the host Gp34 (“prion protein”) as well as other free proteins separated from infectivity. These CJD purifications are better than those previously attained in scrapie, and may be useful for further studies of non-host protein and nucleic acid species. The data are consistent with the hypothesis that CJD-like agents are composed of nucleic acid-protein complexes.