title: Protein stability, conformational change and binding mechanism of human serum albumin upon binding of embelin and its role in disease control.
abstract: Here, we present the inclusive binding mode of phytochemical embelin, an anticancer drug with human serum albumin (HSA) established under physiological condition. Also, to understand the pharmacological role of embelin molecule, here, we have studied the anti-cancer activity of embelin on human cervical cancer cell line (HeLa cell line), which revealed that embelin showed dose dependent inhibition in the growth of cancer cells and also induces 26.3% of apoptosis at an IC50 value of 29μM. Further, embelin was titrated with HSA and the fluorescence emission quenching of HSA due to the formation of the HSA-embelin complex was observed. The binding constant of this complex is 5.9±.01×10(4)M(-1) and the number of bound embelin molecules is approximately 1.0. Consequently, molecular displacement and computational docking experiments show that the embelin is binding to subdomain IB to HSA. Further evidence from microTOF-Q mass spectrometry showed an increase in mass from 66,563Da to 66,857Da observed for free HSA and HSA+embelin complex, signifying that there is robust binding of embelin with HSA. In addition, the variations of HSA secondary structural elements in presence of embelin were confirmed by circular dichroism which indicates partial unfolding of protein. Furthermore, the transmission electron micrographs established that complex formation leads to aggregation of HSA plus embelin. Molecular dynamics simulations revealed that the stability of the HSA-embelin complexes and results suggests that at around 3500ps the complex reaches equilibration state which clearly contributes to the understanding of the stability of the HSA-embelin complexes.
authors.affiliation: Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, India.
authors.forename: Daniel Pushparaju
authors.initials: DP
authors.lastname: Yeggoni
authors.affiliation: National Institute of Animal Biotechnology, Axis Clinicals Building, Miyapur, Hyderabad 500049, India.
authors.forename: Aparna
authors.initials: A
authors.lastname: Rachamallu
authors.affiliation: Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, India. Electronic address: srgsl@uohyd.ernet.in.
authors.forename: Rajagopal
authors.initials: R
authors.lastname: Subramanyam
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D016227:Benzoquinones / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D001665:Binding Sites; D002942:Circular Dichroism; D006367:HeLa Cells; D006801:Humans; D046529:Microscopy, Electron, Transmission; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D021241:Spectrometry, Mass, Electrospray Ionization; D013816:Thermodynamics
chemical_list: D016227:Benzoquinones; D012709:Serum Albumin; C010945:embelin
keywords: Free energy; MicroTOF-Q; Molecular docking; Molecular dynamics simulations; Protein conformation; Protein–ligand interaction