Characterization of Aptamer-Protein Complexes by X-ray Crystallography and Alternative Approaches

Abstract

Aptamers are oligonucleotide ligands, either RNA or ssDNA, selected for high-affinity binding to molecular targets, such as small organic molecules, proteins or whole microorganisms. While reports of new aptamers are numerous, characterization of their specific interaction is often restricted to the affinity of binding (KD). Over the years, crystal structures of aptamer-protein complexes have only scarcely become available. Here we describe some relevant technical issues about the process of crystallizing aptamer-protein complexes and highlight some biochemical details on the molecular basis of selected aptamer-protein interactions. In addition, alternative experimental and computational approaches are discussed to study aptamer-protein interactions.

Light-scattering-based analysis of biomolecular interactions

“…No evidence for self-association was found; the hetero-association data proved the availability of exactly two equivalent and independent binding sites on SA for APT, K D  = 190 nM. The latter result suggests that SA acts as a dimer of dimers where APT may bind to the cleft site, as confirmed by mass spectroscopy (Ruigrok et al 2012).

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Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer

“…The challenges surrounding the solution of crystal structures of aptamer-protein complexes were reviewed (8). Although a number of RNA aptamer/protein complex structures have been solved, only the thrombin-aptamer (9-12) and Von Willebrand Factoraptamer complexes (13) have been solved for a DNA aptamer.…”

The Bound Structures of 17β‐Estradiol‐Binding Aptamers

“…Previous research has demonstrated the extraordinary potential of sensor devices constructed from aptamers, single‐stranded DNA or RNA molecules attached to nanoparticles or nanotubes . These “aptasensors” combine the unique optical and electrical properties of the nanomaterials with the ability of aptamers to bind with high affinity and specificity to predefined targets . Targets include specific proteins and small molecules like 17β‐estradiol (E2).…”

“…To engineer a reliable sensor for the selective and sensitive detection of molecules like E2, it is advantageous to have a thorough understanding of aptamer binding properties at the molecular level. Crystal structures of aptamer‐target complexes provide detailed information on molecular interactions, and thus a thorough understanding of binding . Structures providing the coordinates of all atoms may be obtained by techniques such as X‐ray crystallography, and nuclear magnetic resonance .…”

“…Crystal structures of aptamer‐target complexes provide detailed information on molecular interactions, and thus a thorough understanding of binding . Structures providing the coordinates of all atoms may be obtained by techniques such as X‐ray crystallography, and nuclear magnetic resonance . However, obtaining crystal structures of these complexes has proven difficult, and only handfuls are available .…”