Borukhov S I, Kostrov S V, Izotova L S, Orlova M N, Strongin A Ia
Biokhimiia. 1988 Feb;53(2):332-40.
Using SDS-PAAG electrophoresis, gel-permeation HPLC and immunoblotting, it was demonstrated that homogeneous preparations of human leukocyte interferons (alpha-INF)-A, -N and -I1 obtained from the biomass of the corresponding producer strains (Pseudomonas sp.) contained several oligomeric forms produced by way of S-S intermolecular cross-linkage and making up to 10-15%, 4-7% and 2-5% of the total monomeric form content in the protein preparations. Immunologic testing with the use of MAB NK-2 and [125I]NK-2 showed that the oligomeric forms of alpha-INF-A, -N and -I1 were present in the protein preparations at all purification stages and seemed to be formed at early steps of interferon synthesis in the cell. The effects of limited proteolysis as well as of acid, alkaline and thermodenaturation on the aggregation and oligomerization of alpha-INF-A were studied. SDS-PAAG electrophoresis performed in the absence of the reducing agents showed that upon denaturation of 10% TCA, the amount of the oligomeric forms in the preparations of homogeneous and especially partly proteolytic INF was significantly increased. The causes and the putative mechanisms of aggregation and oligomerization of INF are discussed.
利用SDS-PAAG电泳、凝胶渗透高效液相色谱法和免疫印迹法，证实从相应生产菌株（假单胞菌属）的生物质中获得的人白细胞干扰素（α-INF）-A、-N和-I1的均一制剂含有通过S-S分子间交联产生的几种寡聚形式，分别占蛋白质制剂中总单体形式含量的10 - 15%、4 - 7%和2 - 5%。使用单克隆抗体NK-2和[125I]NK-2进行的免疫学检测表明，α-INF-A、-N和-I1的寡聚形式在所有纯化阶段的蛋白质制剂中均存在，并且似乎是在细胞内干扰素合成的早期步骤形成的。研究了有限蛋白酶解以及酸、碱和热变性对α-INF-A聚集和寡聚化的影响。在不存在还原剂的情况下进行的SDS-PAAG电泳表明，经10%三氯乙酸变性后，均一的尤其是部分蛋白酶解的INF制剂中寡聚形式的量显著增加。讨论了INF聚集和寡聚化的原因及可能机制。
