Exploration of the formation and structure characteristics of a miR-92a promoter G-quadruplex by ESI-MS and CD.

Combining electrospray ionization mass spectrometry (ESI-MS) with circular dichroism (CD) spectroscopy, a G-rich sequence from miR-92a promoter region was discovered to form a parallel G-quadruplex structures in KCl or NHOAc solution. In case of high concentration of NHOAc, the ESI mass spectra showed peaks of a dimeric G-quadruplex structure with 4 ammonium ions. Meanwhile, palmatine, a natural alkaloid, was screened by ESI-MS to bind with the miR-92a G-quadruplex affinitively. The variable temperature experiment of CD also proved that high concentration of NHOAc or palmatine could promote the stability of the dimeric G-quadruplex structure. To get the specific characteristics of the miR-92a G-quadruplex structure, systematic mutations of guanine were tested. Based on the number of NH or ligands, the important guanines involved in the G-quadruplex could be determined. Considering the importance of involved guanines and the number of G-quartets, we speculated an interlocked dimeric parallel G-quadruplex as a possible conformation of the miR-92a promoter G-quadruplex. All results obtained from ESI-MS and CD illustrate structure characteristics of the miR-92a G-quadruplex, which is a promising method for preliminary structural analysis of G-quadruplexes. Besides, this study also provides a strategy for regulating the functions of microRNA by exploring and targeting higher-order structures of miRNAs.