Hohman R J, Rhee S G, Stadtman E R
Proc Natl Acad Sci U S A. 1980 Dec;77(12):7410-4. doi: 10.1073/pnas.77.12.7410.
Sheep antibodies directed against an AMP-bovine serum albumin conjugate exhibit highly specific binding toward AMP. These antibodies bind to the AMP moiety of adenylylated glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] from Escherichia coli and to no other antigenic determinant on the protein. E coli glutamine synthetase can exist in variously modified (isomeric) forms that differ with respect to the number (0-12) and the distribution of identically adenylylated subunits [Ciardi, J. E., Cimino, F. & Stadtman, E. R. (1973) biochemistry 12, 4321-4330]. Using this enzyme, together with the AMP-specific antibodies, we have investigated the effects of the total concentration, population density, and topographical distribution of multiple identical antigenic determinants on the antigen-antibody interaction. Stopped flow fluorescence measurements show that the rate and extent of initial binding of the antibodies to the antigen are a function of the total concentration of AMP groups and are independent of the number of AMP groups der dodecamer. However, the rate of lattice formation increases with increasing epitope density, and the maximal amount of glutamine synthetase precipitated is directly proportional to the average number of adenylylated subunits per dodecamer. The data suggest that partially adenylylated enzyme preparations are composed of subpopulations of glutamine synthetase molecules that differ in their tendency to form precipitable aggregates, due presumably to differences in the topographical distribution of antigenic determinants on the surface of the enzyme. The enzyme species that form soluble immune complexes do so possibly due to intramolecular crosslinkage of the bivalent antibodies with adenylylated subunits to the exclusion of intermolecular crosslinkage.
针对AMP-牛血清白蛋白偶联物的绵羊抗体对AMP表现出高度特异性结合。这些抗体与来自大肠杆菌的腺苷酸化谷氨酰胺合成酶[L-谷氨酸：氨连接酶（ADP形成），EC 6.3.1.2]的AMP部分结合，而不与该蛋白质上的其他抗原决定簇结合。大肠杆菌谷氨酰胺合成酶可以以各种修饰（异构）形式存在，这些形式在相同腺苷酸化亚基的数量（0-12）和分布方面有所不同[恰尔迪，J.E.，奇米诺，F.和斯塔特曼，E.R.（1973年）《生物化学》12卷，4321-4330页]。利用这种酶以及AMP特异性抗体，我们研究了多个相同抗原决定簇的总浓度、群体密度和拓扑分布对抗原-抗体相互作用的影响。停流荧光测量表明，抗体与抗原初始结合的速率和程度是AMP基团总浓度的函数，并且与十二聚体上AMP基团的数量无关。然而，晶格形成的速率随着表位密度的增加而增加，并且沉淀的谷氨酰胺合成酶的最大量与每个十二聚体腺苷酸化亚基的平均数量成正比。数据表明，部分腺苷酸化的酶制剂由谷氨酰胺合成酶分子的亚群组成，这些亚群形成可沉淀聚集体的倾向不同，这可能是由于酶表面抗原决定簇的拓扑分布不同所致。形成可溶性免疫复合物的酶种类可能是由于二价抗体与腺苷酸化亚基的分子内交联，从而排除了分子间交联。
