Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
J Biosci. 2013 Dec;38(5):845-55. doi: 10.1007/s12038-013-9381-0.
Internal mobility of the two domain molecule of ribosome recycling factor (RRF) is known to be important for its action. Mycobacterium tuberculosis RRF does not complement E. coli for its deficiency of RRF (in the presence of E. coli EF-G alone). Crystal structure had revealed higher rigidity of the M. tuberculosis RRF due to the presence of additional salt bridges between domains. Two inter-domain salt bridges and one between the linker region and the domain containing C-terminal residues were disrupted by appropriate mutations. Except for a C-terminal deletion mutant, all mutants showed RRF activity in E. coli when M. tuberculosis EF-G was also co-expressed. The crystal structures of the point mutants, that of the C-terminal deletion mutant and that of the protein grown in the presence of a detergent, were determined. The increased mobility resulting from the disruption of the salt bridge involving the hinge region allows the appropriate mutant to weakly complement E. coli for its deficiency of RRF even in the absence of simultaneous expression of the mycobacterial EF-G. The loss of activity of the C-terminal deletion mutant appears to be partly due to the rigidification of the molecule consequent to changes in the hinge region.
核糖体回收因子（RRF）的两个结构域分子的内部迁移性对于其功能很重要。结核分枝杆菌 RRF 不能在仅存在大肠杆菌 EF-G 的情况下补充大肠杆菌 RRF 的缺乏。晶体结构显示，由于两个结构域之间存在额外的盐桥，结核分枝杆菌 RRF 的刚性更高。通过适当的突变破坏了两个结构域之间的两个盐桥和连接区与包含 C 末端残基的结构域之间的一个盐桥。除了 C 末端缺失突变体之外，当同时表达结核分枝杆菌 EF-G 时，所有突变体在大肠杆菌中都显示出 RRF 活性。确定了点突变体、C 末端缺失突变体和在去污剂存在下生长的蛋白质的晶体结构。涉及铰链区的盐桥的破坏导致的迁移性增加，使适当的突变体即使在没有同时表达分枝杆菌 EF-G 的情况下，也能弱互补大肠杆菌 RRF 的缺乏。C 末端缺失突变体活性的丧失似乎部分是由于铰链区的变化导致分子的僵化。
