Westhoff C M, Lopez O, Goebel P, Carlson L, Carlson R R, Wagner F W, Schuster S M, Wylie D E
School of Biological Sciences, University of Nebraska, Lincoln.
Proteins. 1999 Nov 15;37(3):429-40. doi: 10.1002/(sici)1097-0134(19991115)37:3<429::aid-prot10>3.0.co;2-p.
Monoclonal antibodies (mAb) specific for mercuric ions were isolated from BALB/c mice injected with a mercury-containing, hapten-carrier complex. The antibodies reacted by enzyme-linked immunosorbent assay with bovine serum albumin-glutathione-mercuric chloride (BSA-GSH-HgCl) but not with BSA-GSH without mercury. Nucleotide sequences from polymerase chain reaction products encoding six of the antibody heavy-chain variable regions and seven light-chain variable regions revealed that all the antibodies contained an unpaired cysteine residue in one hypervariable region, which is unusual for murine antibodies. Mutagenesis of the cysteine to either tyrosine or serine in one of the Hg-binding antibodies, mAb 4A10, eliminated mercury binding. However, of two influenza-specific antibodies that contain cysteine residues at the same position as mAb 4A10, one reacted with mercury, although not so strongly as 4A10, whereas the other did not react at all. These results suggested that, in addition to an unpaired cysteine, there are other structural features, not yet identified, that are important for creating an appropriate environment for mercury binding. The antibodies described here could be useful for investigating mechanisms of metal-protein interactions and for characterizing antibody responses to structurally simple haptens.
从注射含汞半抗原 - 载体复合物的BALB/c小鼠中分离出对汞离子具有特异性的单克隆抗体（mAb）。这些抗体通过酶联免疫吸附测定法与牛血清白蛋白 - 谷胱甘肽 - 氯化汞（BSA - GSH - HgCl）发生反应，但不与不含汞的BSA - GSH发生反应。对编码六个抗体重链可变区和七个轻链可变区的聚合酶链反应产物的核苷酸序列分析表明，所有抗体在一个高变区都含有一个未配对的半胱氨酸残基，这在鼠源抗体中是不常见的。在一种汞结合抗体mAb 4A10中，将半胱氨酸突变为酪氨酸或丝氨酸会消除汞结合。然而，在两种与mAb 4A10在相同位置含有半胱氨酸残基的流感特异性抗体中，一种与汞发生反应，尽管反应强度不如4A10，而另一种则根本不发生反应。这些结果表明，除了未配对的半胱氨酸外，还有其他尚未确定的结构特征，对于形成适合汞结合的环境很重要。本文所述的抗体可用于研究金属 - 蛋白质相互作用的机制以及表征对结构简单半抗原的抗体反应。
