Moffat J G, Donly B C, McCaughan K K, Tate W P
Department of Biochemistry, University of Otago, Dunedin, New Zealand.
Eur J Biochem. 1993 Apr 15;213(2):749-56. doi: 10.1111/j.1432-1033.1993.tb17816.x.
Chimeras between Escherichia coli release factors RF-1 and RF-2 have been constructed to study the role of the release factors in termination, in particular whether each possesses specific domains for recognition of the stop codon, and for facilitating peptidyl-tRNA hydrolysis. One hybrid factor showed normal codon-recognition activity but was defective in its ability to facilitate hydrolysis. Overexpression of this protein was toxic to the cell. Conversely, another hybrid factor showed complete loss of codon recognition but retained some hydrolysis activity. These two functional activities of the release factors were not localised in domains within either the amino-terminal or carboxy-terminal halves of the primary sequence as previously predicted. Evidence from the activities of the hybrid proteins and from earlier studies suggests that a combination of residues from the beginning and middle of the sequence, including a region of very high sequence conservation, contribute to the hydrolysis domain, whereas residues from both the amino-terminal and carboxy-terminal halves of the molecule are important for the codon recognition domain.
构建了大肠杆菌释放因子RF-1和RF-2之间的嵌合体，以研究释放因子在终止过程中的作用，特别是它们是否各自拥有识别终止密码子以及促进肽基-tRNA水解的特定结构域。一种杂合因子显示出正常的密码子识别活性，但在促进水解的能力方面存在缺陷。这种蛋白质的过表达对细胞有毒性。相反，另一种杂合因子显示出完全丧失密码子识别能力，但保留了一些水解活性。释放因子的这两种功能活性并不像先前预测的那样定位于一级序列的氨基末端或羧基末端两半的结构域中。来自杂合蛋白活性的证据以及早期研究表明，序列起始和中间部分的残基组合，包括一个序列高度保守的区域，对水解结构域有贡献，而分子氨基末端和羧基末端两半的残基对密码子识别结构域都很重要。
