Formation and Properties of G-quadruplex Formed from Kras Promoter by ESI-MS

G-quadruplexes are special secondary structures of nucleic acids that widely exist in human genome, and it has been suggested that these structures could be involved in many cellular processes, such as gene transcription, cell growth and apoptosis. DNA or RNA that fold into G-quadruplexes are usually guanine-rich sequences, such as human telomere sequence and the promoter regions of certain oncogenes. Since G-quadruplexes represent potentially important targets in cancer relief and drug development, information about the formation, properties, recognition and biological behaviors of G-quadruplexes may provide useful guidance on tumor prevention and treatment. The Kras gene, which is one of the most important cancer genes, has been found to be closely related to human hepatocellular carcinoma, pancreatic cancer, and so on. Recently more and more studies focused on a nuclease hypersensitive element (NHE) located in the promoter region of Kras, which was upstream of the transcription start site. The G-rich strand of NHE was found to form a G-quadruplex suspected to be involved in transcription regulations. So the G-rich strand d (GGGAAGAGGGAAGAGGGGGAGG) of NHE was chosen as the target. Electrospray mass spectrometry (ESI-MS) and circular dichroism (CD) were used to investigate the formation and conformation characteristics of this G-quadruplex in this paper. The results showed that the native type of Kras sequence (KS1) could fold into stable G-quadruplex structures with three tetrads. Furthermore, the mutant experiment was performed to investigate the contributions of different guanines (G) of KS1 to the formation of G-quadruplexes. The results indicat that the sixth G (from 5’ to 3’) of KS1 don’t involve in the formation of G-tetrad, and when it is replaced by thymine (T), the conformation and stability of G-quadruplexes are changed little. On the contrary, the thirteenth, eighteenth and nineteenth G (from 5’ to 3’), show big differences to the formation of G-tetrads. The mutant would cause the conformation of G-quadruplexes transformed into anti-parallel motif and the stability decreased obviously. Since it is still unknown for the NMR or X-ray structure of Kras G-quadruplex so far, the study will be contribute to the structure exploration of Kras G-quadruplex, and will be help in cancer relief as well.