Kornacki J A, West A H, Firshein W
Plasmid. 1984 Jan;11(1):48-57. doi: 10.1016/0147-619x(84)90006-4.
The broad host range plasmid RK2 has previously been found to contain three separate regions of the genome involved in replication and maintenance in Escherichia coli (C. M. Thomas, R. Meyer, D. R. Helinski, 1980, J. Bacteriol. 141, 213-222). They include the origin of replication (oriRK2) and the trfA region which encodes a trans-acting function required for replication. The third region (trfB), although not essential for replication, supplies a function involved in the maintenance of plasmid RK2. Using the maxicell system of labeling plasmid-specific proteins, we have identified all of the proteins encoded by two miniplasmid derivatives of RK2 which contain only the regions oriRK2, trfA, and trfB. To determine which region specifies each protein, RK2/mini-ColE1 hybrid plasmids were used which contain various restriction fragments of the mini-RK2 replicon. The trfA region appears to encode three proteins designated A1 (39,000 MW), A2 (31,000 MW), and A3 (14,000 MW). Analysis of proteins synthesized by plasmids containing deleted forms of the trfA region indicates that the A2 protein is the essential trfA-encoded replication protein of plasmid RK2. The proteins A1 and A3 may be the products specified by the genes tra3 (involved in transmissibility) and kilB1 (involved in host-cell viability) which also map in the trfA region. The trfB region specifies two proteins designated B1 (36,000 MW) and B2 (30,000 MW). These may be the products of the two kil-override (kor) genes located in the trfB region which have been implicated in plasmid maintenance.
此前已发现广宿主范围质粒RK2的基因组包含三个独立区域，参与在大肠杆菌中的复制和维持（C.M.托马斯、R.迈耶、D.R.赫林斯基，1980年，《细菌学杂志》141卷，213 - 222页）。它们包括复制起点（oriRK2）和编码复制所需反式作用功能的trfA区域。第三个区域（trfB）虽然对复制不是必需的，但提供了参与质粒RK2维持的功能。利用标记质粒特异性蛋白质的大细胞系统，我们鉴定了RK2的两种微型质粒衍生物所编码的所有蛋白质，这两种衍生物仅包含oriRK2、trfA和trfB区域。为了确定哪个区域指定每种蛋白质，使用了RK2/微型ColE1杂交质粒，其包含微型RK2复制子的各种限制片段。trfA区域似乎编码三种蛋白质，命名为A1（39,000道尔顿）、A2（31,000道尔顿）和A3（14,000道尔顿）。对含有trfA区域缺失形式的质粒合成的蛋白质分析表明，A2蛋白是质粒RK2中trfA编码的必需复制蛋白。蛋白质A1和A3可能是由也位于trfA区域的tra3基因（参与可转移性）和kilB1基因（参与宿主细胞活力）指定的产物。trfB区域指定两种蛋白质，命名为B1（36,000道尔顿）和B2（30,000道尔顿）。这些可能是位于trfB区域的两个kil - 抑制（kor）基因的产物，它们与质粒维持有关。
