Bis-benzimidazole family molecules are potential anticancer drugs. The dye Hoechst- 33258 is a bis-benzimidazole derivative with an N-methylpiperazine and a phenyl group at the end, and it has been widely used as a fluorescent cytological stain for DNA. It is a long, flexible molecule with a positively charged end and a number of proton donor and acceptor groups suitable for possible hydrogen bonds formation. Hoechst-33258 has been found to self-aggregate because of its planar aromatic hydrophobic structure, that also makes it able to bind in the minor groove of DNA. We have performed a thermodynamic (spectrofluorimetric titrations and quenching measurements) and kinetic (T-jump technique) analysis of Hoechst-33258 self-aggregation.The obtained results show that a dimerization process is coupled with an isomerization reaction (Scheme (1)-(2)): (1) 2D = D2; (2) D= D'. The values of the equilibrium constants obtained from the fluorimetric titrations, quenching measurements and the T-jump kinetic curves are very similar, thus indicating that the three techniques have measured the same processes. The accessible fraction of Hoechst-33258 molecules that we have calculated from quenching measurements reveals that almost 50% of dye’s surface is inaccessible to the quencher (NaI) and this is consistent with the presence of a considerable amount of dimer. The nature of the dye molecule and the results that we have obtained, enable us to hypothesize that isomerization may consist in the slow rotation of the central bond, which exhibits partial characteristic of double bond. One of the two isomers can be stabilized by hydrogen bonds formed through the intermediacy of a water molecule.

Studies on self-Aggregation and Isomerization of the DNA Groove Binder Hoechst-33258

BIVER, TARITA;SECCO, FERNANDO;VENTURINI, MARCELLA
2013

Abstract

Bis-benzimidazole family molecules are potential anticancer drugs. The dye Hoechst- 33258 is a bis-benzimidazole derivative with an N-methylpiperazine and a phenyl group at the end, and it has been widely used as a fluorescent cytological stain for DNA. It is a long, flexible molecule with a positively charged end and a number of proton donor and acceptor groups suitable for possible hydrogen bonds formation. Hoechst-33258 has been found to self-aggregate because of its planar aromatic hydrophobic structure, that also makes it able to bind in the minor groove of DNA. We have performed a thermodynamic (spectrofluorimetric titrations and quenching measurements) and kinetic (T-jump technique) analysis of Hoechst-33258 self-aggregation.The obtained results show that a dimerization process is coupled with an isomerization reaction (Scheme (1)-(2)): (1) 2D = D2; (2) D= D'. The values of the equilibrium constants obtained from the fluorimetric titrations, quenching measurements and the T-jump kinetic curves are very similar, thus indicating that the three techniques have measured the same processes. The accessible fraction of Hoechst-33258 molecules that we have calculated from quenching measurements reveals that almost 50% of dye’s surface is inaccessible to the quencher (NaI) and this is consistent with the presence of a considerable amount of dimer. The nature of the dye molecule and the results that we have obtained, enable us to hypothesize that isomerization may consist in the slow rotation of the central bond, which exhibits partial characteristic of double bond. One of the two isomers can be stabilized by hydrogen bonds formed through the intermediacy of a water molecule.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11568/237952
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