Chambers D A, Zubay G
Proc Natl Acad Sci U S A. 1969 May;63(1):118-22. doi: 10.1073/pnas.63.1.118.
A cell-free system allowing for synthesis of beta-galactosidase enzymatic activity has been developed. This system requires DNA containing the beta-galactosidase gene, a cell-free extract of Escherichia coli bacteria, and the low-molecular-weight components necessary for transcription of the DNA and translation of the resulting messenger RNA. Such a system is useful for studying enzyme synthesis, as well as its regulation. The gene for beta-galactosidase is part of the lac operon whose expression is under the control of the lac repressor. In whole cells the lac repressor inhibits almost all of the gene expression for beta-galactosidase. In the cell-free system, we had previously been able to repress about half the gene expression. Adding cyclic adenosine 3'5'-monophosphate to the cell-free system improved the yield of beta-galactosidase enzymatic activity by 8 to 30 times and the efficiency of repression from 50 to 95 per cent.
已开发出一种无细胞系统，可用于合成β-半乳糖苷酶的酶活性。该系统需要含有β-半乳糖苷酶基因的DNA、大肠杆菌细菌的无细胞提取物，以及DNA转录和由此产生的信使RNA翻译所需的低分子量成分。这样的系统对于研究酶的合成及其调控很有用。β-半乳糖苷酶基因是乳糖操纵子的一部分，其表达受乳糖阻遏物的控制。在完整细胞中，乳糖阻遏物几乎抑制了β-半乳糖苷酶的所有基因表达。在无细胞系统中，我们之前能够抑制大约一半的基因表达。向无细胞系统中添加环腺苷3',5'-单磷酸可使β-半乳糖苷酶的酶活性产量提高8至30倍，阻遏效率从50%提高到95%。
