Investigation of the interactions between Human Telomeric G-Quadruplex and BMVC derivatives
Date Issued
2014
Date
2014
Author(s)
Chen, Cheng-Wei
Abstract
Abstract
Human telomeric DNA consist of tandem repeats of the hexanucleotide d(TTAGGG)n which could form G-quadruplex structure under physiological conditions. Importantly, the formation of G-quadruplexes has been shown to inhibit the activity of the telomerase which is aberrant in cancer cell. Thus, design a G-quadruplex ligand which can target and further stabilize G-quadruplex structures has becoming a potential anticancer strategy.
A novel method is necessary to examine the G-quadruplex ligands with emerging number of small molecules reported in recent years. In general, thermal melting analysis is a typical method used, in which the unfolding of G-quadruplex is monitored by changes in spectroscopic signal. By addition of G-quadruplex ligand, one can obtain the increment in melting temperature (Tm) of G-quadruplexes. However, the major issue of thermal melting analysis method is the dependence on large temperature change. At high temperature, some compounds would induce high-order structure of G-quadruplexes and melting analysis result may not be correspondent to the ability of G-quadruplex ligand at the physiological temperature. Moreover, thermal melting analysis cannot measure additional reliable thermodynamic data, e.g. binding enthalpy, to further design or modify present G-quadruplex ligand. Therefore, we here introduce isothermal titration calorimetry (ITC) to investigate the thermodynamic binding feature of our G-quadruplex ligand, BMVC derivatives bind to human telomeric G-quadruplex, Tel23 dTAG3(T2AG3)3 under isothermal condition. In addition, we used hybridization competition ITC Assay to further evaluate ligand’s ability to stabilize G4 structures.
In order to study and compare G4 ligands, we have selected four G-quadruplex ligands with similar structure, which can be compared by two aspects: Core and Side Chain. The Cores of two isomers are different, one is para-pyridium structure of BMVC Core and the other is ortho-pyridium structure o-BMVC Core. And, two different Side Chain are used, one is hydrophobic four carbon chain, 4C, and the another is hydrophilic triethylene glycol chain, 6C2O. We use ITC to estimate four ligand’s thermodynamic binding parameter. From ITC result, we found out the relationship between G-quadruplex ligand structure and its binding mode. The binding thermodynamic feature of G-quadruplex ligand, for ortho-pyridium structure o-BMVC Core is more enthalpy driven and for para-pyridium structure of BMVC Core is more entropy driven. The difference of side chain has minor effect on thermodynamic feature. It shows core structure of G-quadruplex ligand is the major drive for their binding stability efficiency. Moreover, we further obtain the similar binding heat capacity value for similar core structure that agrees this result.
However, what is the minor effect on the different side chain of ligand ? We further discuss functional side chain’s effect. We have previously reported that BMVC - 6C2O, which constitutes para-pyridium structure of BMVC, can induce structural conversion from nonparallel to parallel. From the previous result, we have known that BMVC-derivatives with different Core have different thermodynamic binding driving force. Thus, we question if the core of ligand governs the binding force, does o-BMVC- 6C2O, which contains different core structure, induce such a structure change from nonparallel G4 to parallel G4 ? If so, do they have different conversion rate and parallel G4 stability? Our data shows that both ligand with 6C2O unit can make structure conversion from nonparallel to parallel. However, conversion rate and stability have major difference which presumably results from the core structure differences.
In conclusion, base on the isothermal titration calorimetry (ITC) assay which provides thermodynamic parameter for ligand-G-quadruplex binding and the results offer thermodynamic information of the G-quadruplex ligand structure with the additional kinetic study of structural conversion, these information on ligand-G4 stability can give more reliable way to design better and potential G-quadruplex ligand for anticancer drug design.
Subjects
人類端粒G-四股結構
G-四股結構穩定劑
恆溫滴定測焓儀
解旋溫度
熱力學結合參數
SDGs
Type
thesis
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