Patrick D R, Zhang K, Defeo-Jones D, Vuocolo G R, Maigetter R Z, Sardana M K, Oliff A, Heimbrook D C
Department of Cancer Research, Merck Sharp and Dohme Research Laboratories, West Point, Pennsylvania 19486.
J Biol Chem. 1992 Apr 5;267(10):6910-5.
Human papillomaviruses (HPVs) are the etiologic agents responsible for genital warts and are contributing factors in the pathogenesis of human cervical cancer. The HPV E7 gene is transcriptionally active in these diseases and has been shown to transform mammalian cells in vitro. We have expressed and purified the HPV-16 E7 gene product in Escherichia coli. The isolated E7 protein contains zinc in a 1:1 molar ratio. X-ray absorption fine structure studies demonstrated that the zinc is coordinated by 4 sulfur ligands. We sequentially derivatized the E7 cysteines to differentiate between solvent-exposed, metal-bound, and disulfide-associated cysteines. Our results demonstrate that Cys24 and Cys68 are accessible to solvent, while cysteines in the two conserved Cys-X-X-Cys motifs are likely involved in binding zinc. We observed no evidence for the existence of disulfide bonds in recombinant E7 protein under the conditions tested.
人乳头瘤病毒（HPV）是引起尖锐湿疣的病原体，也是人类宫颈癌发病机制中的促成因素。HPV E7基因在这些疾病中具有转录活性，并且已证明在体外可转化哺乳动物细胞。我们已在大肠杆菌中表达并纯化了HPV-16 E7基因产物。分离出的E7蛋白含有摩尔比为1:1的锌。X射线吸收精细结构研究表明，锌由4个硫配体配位。我们依次对E7半胱氨酸进行衍生化，以区分溶剂暴露型、金属结合型和二硫键相关型半胱氨酸。我们的结果表明，Cys24和Cys68可与溶剂接触，而两个保守的Cys-X-X-Cys基序中的半胱氨酸可能参与锌的结合。在测试条件下，我们未观察到重组E7蛋白中存在二硫键的证据。
