A Biophysical Study of the G-Quadruplex-Insulin Interaction

Abstract

The formation of guanine (G)-quadruplex structures in the guanine-rich tandem repeats of the insulin-linked polymorphic region (ILPR) have potential effects on transcription of the insulin gene. Recent studies demonstrate that the ILPR G-quadruplexes can bind to insulin. The energetics of the binding between insulin and the G-quadruplexes formed by the most common ILPR repeat sequence have also been characterized in prior work. We have studied the proton transfer involved in the interaction between insulin and this DNA sequence by conducting isothermal titration calorimetry (ITC) experiments in various buffers and analyzing the observed enthalpy change. The transcriptional activity of a number of ILPR repeat sequences, including the consensus sequence, have been measured previously. Transcriptional activity for less common repeats is significantly lower than that of the consensus sequence but can be increased substantially by varying only one or two nucleotides. To determine the potential role of G-quadruplex formation and stability in regulating transcription, we have studied the second and third most common ILPR repeats as well as their variant sequences that exhibit increased transcriptional activity. ITC was used to characterize the energetics of the binding interaction between insulin and each of the four ILPR repeat sequences. The bulk thermodynamic measurements performed at various temperatures from 20 - 37 degrees Celsius provide insight into these biomolecular interactions.