In silico selection of an aptamer to estrogen receptor alpha using computational docking employing estrogen response elements as aptamer-alike molecules.

Rajesh Ahirwar,Smita Nahar,Srinivasan Ramachandran,Souvik Maiti,Pradip Nahar,Shikha Aggarwal

doi:10.1038/srep21285

Abstract

Aptamers, the chemical-antibody substitute to conventional antibodies, are primarily discovered through SELEX technology involving multi-round selections and enrichment. Circumventing conventional methodology, here we report an in silico selection of aptamers to estrogen receptor alpha (ERα) using RNA analogs of human estrogen response elements (EREs). The inverted repeat nature of ERE and the ability to form stable hairpins were used as criteria to obtain aptamer-alike sequences. Near-native RNA analogs of selected single stranded EREs were modelled and their likelihood to emerge as ERα aptamer was examined using AutoDock Vina, HADDOCK and PatchDock docking. These in silico predictions were validated by measuring the thermodynamic parameters of ERα -RNA interactions using isothermal titration calorimetry. Based on the in silico and in vitro results, we selected a candidate RNA (ERaptR4; 5′-GGGGUCAAGGUGACCCC-3′) having a binding constant (Ka) of 1.02 ± 0.1 × 108 M−1 as an ERα-aptamer. Target-specificity of the selected ERaptR4 aptamer was confirmed through cytochemistry and solid-phase immunoassays. Furthermore, stability analyses identified ERaptR4 resistant to serum and RNase A degradation in presence of ERα. Taken together, an efficient ERα-RNA aptamer is identified using a non-SELEX procedure of aptamer selection. The high-affinity and specificity can be utilized in detection of ERα in breast cancer and related diseases.

Highlights

Aptamers, the chemical-antibody substitute to conventional antibodies, are primarily discovered through systematic evolution of ligands by exponential enrichment (SELEX) technology involving multi-round selections and enrichment
To identify an estrogen receptor α (ERα) -targeting aptamer by a non-SELEX procedure, we hypothesize that the estrogen response elements, which are the stretches of B-DNA in the promoter region of the genes regulated by estrogen receptors[18,19], can be utilized to obtain a pool of aptamer-alike sequences for in silico screening
We have developed a non-SELEX method that combines the in vivo chemistry of estrogen response elements (EREs) structure and interactions with computation modelling and molecular docking to identify an ERα binding aptamer

Summary

Introduction

The chemical-antibody substitute to conventional antibodies, are primarily discovered through SELEX technology involving multi-round selections and enrichment. To identify an ERα -targeting aptamer by a non-SELEX procedure, we hypothesize that the estrogen response elements, which are the stretches of B-DNA in the promoter region of the genes regulated by estrogen receptors[18,19], can be utilized to obtain a pool of aptamer-alike sequences for in silico screening. Their inverted repeat nature and potential to interact with ERα in vivo provides a way to mimic these characteristics in an in silico system. Solid-phase assays are performed to demonstrate the antibody-alternative action of the selected ERα -aptamer

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Conclusion