STRUCTURAL-ANALYSIS OF RAUSCHER VIRUS GP70 USING MONOCLONAL-ANTIBODIES - SITES OF ANTIGENICITY AND P15(E) LINKAGE
STRUCTURAL-ANALYSIS OF RAUSCHER VIRUS GP70 USING MONOCLONAL-ANTIBODIES - SITES OF ANTIGENICITY AND P15(E) LINKAGE
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DOI：
10.1016/0042-6822(82)90305-1
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发表时间：
1982-01-01
期刊：
影响因子：
3.7
通讯作者：
ELDER, JH
中科院分区：
文献类型：
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作者：
NIMAN, HL;ELDER, JH
使用命名为PEC-MAP（部分酶切-单克隆抗体沉淀）的技术，生成Rauscher逆转录病毒gp 70上19个小鼠单克隆抗体结合结构域的线性图谱。用8种蛋白水解酶制备的限制性内切酶产生39个gp 70片段。免疫沉淀这些片段的单克隆抗体从51个细胞系允许19个结合位点被定义凭借重叠的片段和差异沉淀模式。然后通过分析各种gp 70片段的表观MW来绘制最易受蛋白水解攻击和抗体结合的位点。该分析揭示了gp 70中的3个高反应性区域，其大约位于gp 70的氨基末端的前2000道尔顿内以及MW 47，000去糖基化gp 70分子的氨基末端的18，000和38，000道尔顿内。针对二硫键连接的p15（E）分子的单克隆抗体用于将其连接位点定位于结构域XVII，估计距离去糖基化gp 70的氨基端34，000 - 38，000道尔顿。这些数据对gp 70的三级结构有一定的限制，并提出了产生白血病MCF重组体的机制。
A linear map of 19 mouse monoclonal antibody-binding domains on Rauscher retroviral gp70 was generated using a technique designated PEC-MAP (partial enzymatic cleavage-monoclonal antibody precipitation). Eight proteolytic enzyme preparations in limited digests were used to produce 39 gp70 fragments. Immune precipitation of these fragments by monoclonal antibodies from 51 cell lines allowed 19 binding sites to be defined by virtue of overlapping fragments and differential precipitation patterns. The sites most accessible to proteolytic attack and antibody binding were then mapped by analyzing the apparent MW of the various gp70 fragments. This analysis revealed 3 hyperreactive regions in gp70 located approximately within the first 2000 daltons of the amino end of gp70 and 18,000 and 38,000 daltons from the amino end of the MW 47,000 deglycosylated gp70 molecule. Monoclonal antibodies directed against the disulfide-linked p15(E) molecule were used to localize its linkage site to Domain XVII, estimated to be 34,000-38,000 daltons from the amino end of deglycosylated gp70. These data place certain constraints on the tertiary structure of gp70 and suggest a mechanism for the generation of leukemogenic MCF recombinants.