title: Electrospray ionization mass spectrometric exploration of the high-affinity binding of three natural alkaloids with the mRNA G-quadruplex in the BCL2 5'-untranslated region.
abstract: BACKGROUND
  The BCL2 gene encodes an integral outer mitochondrial membrane protein (25 kDa) which regulates the apoptotic death of cells. There is a 25-nucleotide G-rich sequence in the 5'-untranslated region (5'-UTR) of the BCL2 mRNA, which can adopt a G-quadruplex structure. Small molecules which could tightly bind to this structure have a potential function in the regulation of the expression of the BCL2 mRNA.
  
  
  METHODS
  The 25-mer oligonucleotide (5'-G(5)CCGUG(4)UG(3)AGCUG(4)-3') was synthesized by TaKaRa Biotechnology Co., Ltd. (TaKaRa, Dalian) with high-performance liquid chromatography (HPLC) purification. Electrospray ionization (ESI) mass spectrometry (MS) was used to probe the binding properties of natural small molecules (P) with the mRNA G-quadruplex in the BCL2 5'-UTR (BCL2Q). Collision-induced dissociation (CID) mass spectrometry and circular dichroism (CD) spectroscopy were performed to evaluate the stabilization of the mRNA G-quadruplex and its complexes.
  
  
  RESULTS
  The results from ESI mass spectra showed that three natural alkaloids (nitidine, palmatine, and jatrorrizine) have high binding affinities to the mRNA G-quadruplex with the binding stoichiometry ranging from 1:1 to 3:1. CID mass spectrometry results revealed that the G-quadruplex-ligand complex lost bases first rather than losing the binding molecules. Increases in the T(m) values of the complexes of the G-quadruplex with the natural alkaloids in the CD melting experiments demonstrated that the three small molecules can stabilize the G-quadruplex structure.
  
  
  CONCLUSIONS
  Three natural small molecules were found to have very high binding affinities to the mRNA G-quadruplex and stabilize this structure. The properties of these alkaloids revealed promising potentials to regulate the expression of the BCL2 protein from the posttranscriptional pathway.
authors: Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
authors: Wei
authors: W
authors: Tan
authors: Gu
authors: G
authors: Yuan
mesh_terms: D000470:Alkaloids / Q000737:chemistry* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D054856:G-Quadruplexes; D019254:Genes, bcl-2; D006801:Humans; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D020506:Untranslated Regions
chemical_list: D000470:Alkaloids; D012333:RNA, Messenger; D020506:Untranslated Regions