Robertson M P, Ellington A D
Department of Chemistry and Institute for Cellular and Molecular Biology, University of Texas at Austin, 78712, USA.
Nat Biotechnol. 1999 Jan;17(1):62-6. doi: 10.1038/5236.
We have selected an allosteric ribozyme ligase from a random sequence population that is activated up to 10,000-fold by oligonucleotide effectors. The ribozyme conforms to a classic two-state model for allostery in which the equilibrium between inactive and active conformers is dramatically altered by the presence of effector ligands. In the presence of the effector the allosteric ribozyme ligase generates templates that can subsequently be amplified using conventional amplification technologies, such as RT-PCR. Thus, the allosteric ribozyme can transduce (or convert) analytes into amplicons. We demonstrate two potential diagnostic applications of the selected allosteric ribozyme ligase: 'counting' short oligonucleotide effectors by RT-PCR, and counting a non-nucleic acid effector, ATP, by ligation.
我们从一个随机序列群体中筛选出了一种变构核酶连接酶，它可被寡核苷酸效应物激活达10000倍。该核酶符合变构作用的经典双态模型，即效应物配体的存在会显著改变无活性和活性构象体之间的平衡。在效应物存在的情况下，变构核酶连接酶会生成模板，随后可使用常规扩增技术（如逆转录聚合酶链反应，RT-PCR）对其进行扩增。因此，变构核酶可将分析物转导（或转化）为扩增子。我们展示了所选变构核酶连接酶的两种潜在诊断应用：通过RT-PCR“计数”短寡核苷酸效应物，以及通过连接反应“计数”非核酸效应物ATP。
