Spherical-Shape Assumption for Protein–Aptamer Complexes Facilitates Prediction of Their Electrophoretic Mobility

Abstract

Supporting information placeholder ABSTRACT: DNA aptamers are single-strand DNA (ssDNA) capable of selectively and tightly binding a target molecule. Capillary electrophoresis-based selection of aptamers for protein targets requires the knowledge of electrophoretic mobilities of protein-aptamer complexes while measuring these mobilities requires having the aptamers. Here we report on breaking this vicious circle. We introduce a mathematical model that allows accurate prediction of proteinaptamer complex mobility while requiring only three easy-to-determine input parameters: the number N of nucleotides in the aptamer, electrophoretic mobility of N-nucleotide-long ssDNA, and a sum molecular weight of the protein-aptamer complex. The model is built upon two simplifying assumptions: a spherical shape of the protein-aptamer complex and a negligible charge of the protein with respect to that of the aptamer. According to this model, the protein-aptamer complex mobility is a linear function of a combination of the three input parameters with empirically determined line's intercept and slope. The intercept and slope were determined using experimental data for seven complexes. The model was then cross-validated with the leave-one-out approach revealing only 2% residual standard deviations for both the slope and the intercept. Such a precise determination of these constants allowed accurate mobility prediction for the excluded complexes with only a 3% maximum deviation from the experimentally determined mobilities. Our results strongly suggest the validity of the spherical-shape assumption for the protein-aptamer complexes when considering complex mobility. The developed model will make it possible to rationally design capillary electrophoresis-based selection of DNA aptamers for protein targets.

Empirical predictor of conditions that support ideal‐filter capillary electrophoresis

“…Two models are applicable to double‐strand DNA and unstructured single‐strand DNA attached to the protein in a single point [8]. The third model predicts μ P–DNA for structured single‐strand DNA (e.g., an aptamer) tightly linked to the protein through a number of bonds [9]. Thus, the values of μ DNA and μ P–DNA can be considered as known and can, thus, serve to determine the range of IFCE‐suitable μ EOF according to Eq.…”

SELEX-based DNA Aptamer Selection: A Perspective from the Advancement of Separation Techniques

“…Belobrodov et al recently reported a mathematical (but empirical) model to predict the  of protein-aptamer complexes. 57 Although  is one of the most important parameters for CE-based selection to determine the partitioning position in an electropherogram, aptamers are needed to measure  To resolve this dilemma, Belobrodov et al established a model for predicting the  of the complex by assuming a spherically shaped complex composed of some known parameters: Nlib,  of free DNA, and a total molecular weight for the complex. The validity of their equation was proven using seven aptamer-protein complexes with only 2% standard deviations.…”

Spatiotemporally Controlled DNA Nanoclamps: Single-Molecule Imaging of Receptor Protein Oligomerization

“…Therefore, how to detect and regulate receptor proteins must be an attractive topic. 10 When it comes to detecting receptor proteins, aptamers, as protein-specific binding units, are widely utilized 11,12 due to their specificity and simplicity of synthesis. 13,14 However, the interaction between aptamer and receptor can result in side effects, such as disturbing the homeostasis of the cell's internal environment because a single receptor protein participates in a variety of cell behaviors.…”

“…When it comes to detecting receptor proteins, aptamers, as protein-specific binding units, are widely utilized , due to their specificity and simplicity of synthesis. , However, the interaction between aptamer and receptor can result in side effects, such as disturbing the homeostasis of the cell’s internal environment because a single receptor protein participates in a variety of cell behaviors . Moreover, it is difficult to avoid the problem of insufficient specificity in single aptamer detection. , Yang et al reported the use of dual aptamers to detect receptor protein oligomerization for the first time, which improved the specificity and reduced the side effects of the probe, providing a platform basis for subsequent work …”