PREPARATION OF PROTEIN CONJUGATES VIA INTERMOLECULAR DISULFIDE BOND FORMATION
PREPARATION OF PROTEIN CONJUGATES VIA INTERMOLECULAR DISULFIDE BOND FORMATION
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DOI：
10.1021/bi00601a022
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发表时间：
1978-01-01
期刊：
影响因子：
2.9
通讯作者：
KOCHOUMIAN, L
中科院分区：
文献类型：
--
作者：
KING, TP;LI, Y;KOCHOUMIAN, L
两种不同蛋白质的缀合物可以通过分子间二硫键交换反应制备，即含有硫醇基团的蛋白质A与含有4-二硫吡啶基基团的蛋白质B反应，产生缀合物并释放4-硫代吡啶酮。硫醇基团可以通过用 3-巯基丙酰亚胺酸甲酯酯或 2-亚氨基四氢噻吩进行脒化引入到蛋白质中，并且可以在 4,4''-二硫代二吡啶存在下用这些相同的试剂将 4-二硫吡啶基团引入到蛋白质中。与已知的亚氨酸酯的不稳定性相比，2-亚氨基硫杂环戊烷在储存时是稳定的；因此，2-亚氨基硫杂环戊烷是比亚氨酸酯更方便的蛋白质修饰试剂。所有反应均可在温和条件下轻松进行，收率良好。用每摩尔平均含有 1-5 个硫醇或二硫代吡啶基团的修饰蛋白制备牛血浆白蛋白与其自身、RNase 或 D-谷氨酸和 D-赖氨酸的共聚物以及绵羊抗体和辣根过氧化物酶的缀合物。这些缀合物主要形成二聚体、三聚体和四聚体。过氧化物酶标记的抗体保留了 80% 以上的酶和抗原结合活性。
Conjugates of 2 unlike proteins can be prepared via the intermolecular disulfide interchange reaction, i.e., protein A containing thiol groups reacts with protein B containing 4-dithiopyridyl groups to yield a conjugate with the release of 4-thiopyridone. Thiol groups can be introduced into proteins on amidination with methyl 3-mercaptopropionimidate ester or 2-iminothiolane, and 4-dithiopyridyl groups can be introduced into proteins with these same reagents in the presence of 4,4''-dithiodipyridine. 2-Iminothiolane is stable on storage in contrast to the known lability of imidate esters; therefore 2-iminothiolane is a more convenient reagent for the modification of proteins than are the imidate esters. All the reactions can be carried out easily under mild conditions in good yields. Conjugates of bovine plasma albumin with itself, RNase or a copolymer of D-glutamic acid and D-lysine and of sheep antibody and horseradish peroxidase were prepared with modified proteins containing an average of 1-5 thiol or dithiopyridyl groups per mol. These conjugates formed mainly dimers, trimers and tetramers. The peroxidase labeled antibody retained more than 80% of its enzymatic and antigenic binding activities.