N-甲基甲卟啉 IX 适体的选择以开发卟啉金属化 DNA 酶。

Selection of Aptamer for N-Methyl Mesoporphyrin IX to Develop Porphyrin Metalation DNAzyme.

机构信息

CAS Key Laboratory of Nano-Bio Interface, Suzhou Institute of Nano-Tech and Nano-Bionics, Chinese Academy of Sciences, Suzhou, China.

School of Nano-Tech and Nano-Bionics, University of Science and Technology of China, Hefei, China.

出版信息

Methods Mol Biol. 2022;2439:15-26. doi: 10.1007/978-1-0716-2047-2_2.

DOI:10.1007/978-1-0716-2047-2_2

PMID:35226312

Abstract

Mesoporphyrin IX (MPIX) contains a planar macrocycle center that can interact with various divalent metal ions through the exposed binding sites, leading to the metalation of MPIX. The DNA aptamers for porphyrin molecules usually display different catalytic functions (termed deoxyribozymes or DNAzymes), which can accelerate such chemical reactions. Inspired by this, an affinity chromatography selection approach was designed for identifying a porphyrin metalation DNAzyme. In our experiment, N-methyl mesoporphyrin IX (NMM), an analog of MPIX, is used as the target molecule, owing to its stable and high fluorescence enhancement after combining with specific oligonucleotides. Our results showed that the selected aptamer Nm1 is capable of binding to NMM with a low micromolar dissociation constant (0.75 ± 0.08 μM) and displays a catalytic activity for MPIX metalation with 3.3-fold rate enhancement. The protocol for isolation of such a porphyrin metalation DNAzyme is described in detail here.

摘要

中卟啉 IX（MPIX）含有一个平面大环中心，通过暴露的结合位点可以与各种二价金属离子相互作用，从而导致 MPIX 的金属化。卟啉分子的 DNA 适体通常表现出不同的催化功能（称为脱氧核酶或 DNA 酶），可以加速这些化学反应。受此启发，设计了一种亲和层析选择方法来鉴定卟啉金属化 DNA 酶。在我们的实验中，N-甲基中卟啉 IX（NMM）用作靶分子，因为它与特定寡核苷酸结合后具有稳定且荧光增强的特性。我们的结果表明，所选择的适体 Nm1 能够以低微摩尔解离常数（0.75±0.08 μM）与 NMM 结合，并表现出对 MPIX 金属化的催化活性，其速率提高了 3.3 倍。本文详细描述了分离这种卟啉金属化 DNA 酶的方案。