RECONSTITUTION OF 7S MOLECULES FROM L AND H POLYPEPTIDE CHAINS OF ANTIBODIES AND GAMMA-GLOBULINS.
RECONSTITUTION OF 7S MOLECULES FROM L AND H POLYPEPTIDE CHAINS OF ANTIBODIES AND GAMMA-GLOBULINS.
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从L和H多肽链的7s分子重构的抗体和γ-球蛋白。
DOI：
10.1084/jem.119.5.789
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发表时间：
1964-05-01
影响因子：
15.3
通讯作者：
EDELMAN, G M
中科院分区：
文献类型：
--
作者：
OLINS, D E;EDELMAN, G M
已发现在适当条件下将7S γ-球蛋白的分离的L和H多肽链混合导致7S分子的重构。将链在0.5 N丙酸中混合，当透析到中性水性缓冲液中时，它们相互作用，以大于30%的产率形成重构材料。该材料的沉降系数为6S ~ 7S，重均分子量为160，000，其抗原结构和电泳特性与7S γ-球蛋白相同。通过在不同类型的链上使用I131和I125标记，结合蔗糖密度梯度中链混合物的超离心，发现7S产物含有与两个L和两个H链的存在一致的比例的两种同位素。用木瓜蛋白酶水解后，重构的材料产生类似于S和F片段的产物。对应于L链的所有同位素都存在于S片段的对应物中;对应于H链部分的同位素存在于两个片段中。从纯化的豚鼠抗体的链重构的活性被认为是主要与7S材料。通过相同的重构技术形成含有兔L链和人H链以及人L链和兔H链的杂交分子。发现天然7S γ-球蛋白的链间二硫键可以断裂和再氧化，重组7S材料的链间二硫键也可以断裂和再氧化。将还原的L和H链混合在丙酸中，用含有Na 2SO 4的中性缓冲液透析，然后在不存在Na 2SO 4的情况下用中性缓冲液透析，形成分子量为160，000的稳定的7S分子，其仅在还原后可解离。
Admixture of separated L and H polypeptide chains of 7S γ-globulins under appropriate conditions has been found to result in the reconstitution of 7S molecules. The chains were mixed in 0.5 N propionic acid and when dialyzed into neutral aqueous buffers interacted to form reconstituted material in greater than 30 per cent yield. This material had sedimentation coefficients of 6S to 7S, a weight average molecular weight of 160,000, and its antigenic structure and electrophoretic properties were the same as those of 7S γ-globulin. By the use of I131 and I125 labels on the different types of chains, combined with ultracentrifugation of chain mixtures in sucrose density gradients, the 7S product was found to contain both isotopes in ratios consistent with the presence of two L and two H chains. After hydrolysis with papain, the reconstituted material yielded products resembling S and F fragments. All of the isotope corresponding to L chains was found in the counterpart of the S fragment; the isotope corresponding to the H chain fraction was present in both fragments. The activity reconstituted from chains of a purified guinea pig antibody to f1 phage was found to be associated mainly with the 7S material. Hybrid molecules containing rabbit L chains and human H chains and of human L chains and rabbit H chains were formed by the same techniques of reconstitution. It was found that the interchain disulfide bonds of native 7S γ-globulins could be broken and reoxidized, as could those of reconstituted 7S material. Reduced L and H chains mixed in propionic acid, dialyzed against neutral buffers containing mercaptan, then against neutral buffers in the absence of mercaptan, formed stable 7S molecules of molecular weight 160,000, which were dissociable only after reduction.