Characterization of doxorubicin binding site and drug induced alteration in the functionally important structural state of oxyhemoglobin

Shahper N. Khan, Barira Islam, Ragothaman Yennamalli, Qamar Zia, Naidu Subbarao, Asad U. Khan

Research output: Contribution to journalArticlepeer-review

23 Citations (Scopus)

Abstract

Doxorubicin (DOX) binding to hemoglobin (Hb) was studied to investigate the drug induced protein dysfunction. The features of anti-tumor drug doxorubicin infused structural perturbation of human Hb were studied by circular dichroism (CD). The mechanism of DOX-Hb binding was elucidated by steady-state and synchronous fluorescence spectroscopy. The Stern-Volmer analysis indicated that the binding of Hb to DOX is characterized by more than one high affinity binding site with the association constants of the order of 105. Hydrophobic probe ANS was employed to elucidate the drug binding site. Binding mode expounded by thermodynamic parameters implied the role of hydrogen bonding, electrostatic and hydrophobic interaction in stabilizing the complex. The molecular distance between donor (Hb) and acceptor (DOX) was calculated according to Förster's theory of energy transfer. Fourier transform infrared (FT-IR) spectroscopy provides an insight to the changes occurring in protein on DOX binding. Treatment of Hb with DOX resulted in a dose dependent fragmentation of protein. The quantitative analysis revealed the release of acid soluble amino groups from the photoexcited Hb-DOX mixture. The free radical mediated degradation was suggested by its rescue on mannitol and superoxide dismutase (SOD) appliance. The loss of protein band further corroborates the concentration dependent Hb fragmentation. The molecular modeling complies with the thermodynamic data of forces involved in DOX binding and depicts its interaction in the proximity of oxygen binding pocket of Hb. Thus, this study enriches our understanding of the interaction dynamics of anticancer drugs to the physiologically important protein Hb.

Original languageEnglish
Pages (from-to)1096-1104
Number of pages9
JournalJournal of Pharmaceutical and Biomedical Analysis
Volume48
Issue number4
Early online date4 Sep 2008
DOIs
Publication statusPublished - 1 Dec 2008
Externally publishedYes

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