Search

Dutasteride is a pharmacologically important drug employed to treat prostate cancer. Alpha-2-macroglobulin (α 2 M) is the primary proteinase inhibitor and is abundant in vertebrate plasma. Previous studies have shown that α 2 M levels were down regulated in prostate cancer. Our results of functional assay shows 50% decrease in the antiproteolytic potential ofα 2 Mupon its interaction with dutasteride. Fluorescence quenching revealed that dutasteride binds with α 2 M via static mechanism, resulting in the formation of dutasteride-α 2 M complex. Synchronous fluorescence studies suggest alteration in the microenvironment around tryptophan residues. Changes in the UV-visible spectra hints at formation of complex between the drug and protein. Secondary structural perturbations in α 2 M are confirmed by circular dichroism studies. Molecular docking discloses the involvement of hydrogen bonding during the interaction process and suggests the site of interaction of dutasteride on α 2 M monomer as Asn173, Lys171, Asp1178, Lys1236, His1182, Lys1177, Ser1180 and Lys1240.Isothermal titration calorimetry affirms the binding process to be spontaneous and exothermic. The results of this study may potentially be important should it be shown that dutasteride interacts with α 2 M under physiological conditions., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Alpha-2-macroglobulin (α 2 M) is large glycoprotein present in the body fluids of vertebrates. It is an antiproteinase that inhibits a broad spectrum of proteases without the direct blockage of the protease active site. Curcumin, a yellow spice commonly used in India and several Asian countries, is reported to have anti-tumor and anti-inflammatory effects because of its antioxidant properties. In the present study, we have explored the interaction of curcumin with α 2 M using various technique such as antiproteinase activity assay, spectroscopy. Changes in the secondary structure of α 2 M following interaction with curcumin was investigated by CD and FT-IR spectroscopy. Thermodynamics of curcumin-α 2 M binding were also analyzed by isothermal titration calorimetry to identify the number of binding sites, changes in enthalpy, entropy and Gibbs free energy changes for this interaction. Thermodynamics parameters reveal that the binding is exothermic in nature. Our results suggest that the binding of curcumin with α 2 M induces a conformational change in the native form of protein that compromises its anti-proteinase activity. This exothermic and spontaneous interaction leads to alteration in the β-sheet content of the protein leading to subtle changes in conformational status of the protein leading possibly to loss in the antiproteinase potential of the inhibitor., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Bicalutamide (BCT), a drug used in the treatment of prostate cancer, antagonises the actions of androgens, at the receptor level, thereby inhibiting the growth of prostate tumours. Alpha-2-macroglobulin (α 2 M), a pan-proteinase inhibitor, inhibits proteinase, regardless of specificity and catalytic mechanism. α 2 M is deficient in patients of advanced prostate cancer with bone metastases. Our studies explored the interaction of BCT with α 2 M and analysed the BCT induced structural alteration to the α 2 M. The result suggests that BCT decreases the antiproteolytic potential and causes structural and functional change in human α 2 M. UV-visible absorption spectroscopy confirms the formation of α 2 M-BCT complex. Fluorescence analysis shows significant quenching in fluorescence intensity of α 2 M upon binding with BCT. Synchronous fluorescence result suggests the interaction of BCT with α 2 M changed the microenvironment around tyrosine residues. Secondary structure of α 2 M also undergoes a slight change upon complexation with the drug as evident by shift in negative ellipticity in far UV CD spectroscopy. FTIR results confirm the alteration in secondary structure of α 2 M upon drug interaction. Molecular docking studies show that BCT bind to a monomer of α 2 M primarily through hydrophobic force. Thermodynamics parameters were determined by isothermal titration calorimetry found that the binding was exothermic in nature., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Hypochlorous acid (HOCl), an active bleaching agent is one of the major oxidant produced by neutrophils under physiological conditions. It is among one of the most potent reactive oxygen species (ROS) which causes oxidation of biomolecules. Treatment of proteins with hypochlorite results in direct oxidative damage to the protein. Alpha-2-macroglobulin (α 2 M) is a major proteinase inhibitor that can inhibit proteinase of any kind regardless of their specificity and catalytic mechanism. The proteinase-antiproteinase balance plays an important role in mediating inflammation associated tissue destruction. In this paper, we intend to study hypochlorite induced modifications in proteinase inhibitor- α 2 M via biophysical techniques such as absorption spectroscopy, fluorescence spectroscopy, circular dichroism (CD), fourier transform infrared spectrometry (FTIR) and isothermal titration calorimetry (ITC). It was found that hypochlorite decreases the anti-proteolytic potential and causes inactivation of sheep α 2 M. It also causes structural and functional change in sheep α 2 M as evident by UV-Visible absorption spectroscopy and fluorescence measurements. Change in secondary structure of α 2 M was confirmed by CD and FTIR. Thermodynamics parameters such as entropy change (ΔS), enthalpy change (ΔH), Gibbs free energy change (ΔG) and the number of binding sites (N) of α 2 M-HOCl binding in solutions were determined by ITC. Moreover, it was found that binding of HOCl with α 2 M was exothermic in nature., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Alpha-2-macroglobulin is a multifunctional, highly abundant, plasma protein which reacts with a wide variety of molecules and drugs including cisplatin. Cisplatin is commonly used anticancer drug widely used for treatment of testicular, bladder, ovarian, head and neck, lung and cervical cancers. This study is designed to examine the interaction of cisplatin with human alpha-2-macroglobulin through various biophysical techniques and drug binding through molecular modeling. Cisplatin alters the function of alpha-2-macroglobulin and the thiolesters are most likely the reactive sites for cisplatin. Our result suggests that cisplatin decreases the antiproteolytic potential and causes structural and functional change in human alpha-2-macroglobulin as evident by absorption and fluorescence spectroscopy. Change in secondary structure of alpha-2-macroglobulin was confirmed by CD and FTIR. Thermodynamics parameters such as entropy (ΔS), enthalpy (ΔH) and Gibb's free energy changes (ΔG) along with number of binding sites (N) of alpha-2-macroglobulin-cisplatin binding in solutions were determined by isothermal titration calorimetry (ITC). It was found that binding of cisplatin with alpha-2-macroglobulin was exothermic in nature. The interaction of drug with alpha-2-macroglobulin in the plasma could lead to structural alterations in the conformational status of alpha-2-macroglobulin resulting in its functional inactivation., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Alpha-2-macroglobulin (α 2 M), a pan-proteinase inhibitor, inhibits a variety of endogenous and exogenous proteinases and constitutes an important part of body's innate defense system. In the present study, we explored how trifluoroethanol (TFE) may modulate the structure, antiproteinase activity and aggregation of α 2 M. TFE was sequentially added over a range of 0-20% (v/v) and the effects induced were studied by activity assay, intrinsic fluorescence, ANS fluorescence, circular dichroism, turbidity assay, Rayleigh scattering measurement and ThT fluorescence measurement. Decrease in activity and increase in fluorescence intensity of α 2 M upon addition of TFE shows structural deviation from the native structure and suggests aggregation of protein upon solvent addition. Increase in turbidity and Rayleigh scattering of modified α 2 M confirms the formation of aggregates. Insignificant ThT fluorescence intensity of TFE treated α 2 M is indicative of amorphous or non-amyloid aggregation. Further, circular dichroism results indicate the changes in secondary structure of native α 2 M as negative ellipticity decreased on addition of the polar solvent to the inhibitor. The turbidometric analysis, Rayleigh scattering, ThT fluorescence intensity of modified α 2 M suggests that the protein might be driven towards non-amyloid or amorphous aggregation. Our studies provide important mechanistic insight how α 2 M undergoes conformational and functional changes when exposed to TFE., (Copyright © 2017 Elsevier B.V. All rights reserved.)

A α2M homologue was isolated from sheep (Ovis aries) blood plasma, using a simple two-step procedure, ammonium sulphate fractionation and gel filtration chromatography. Sheep α2M was found to be a large tetrameric glycoprotein of 630 kDa with monomeric subunit of 133 kDa each. Each subunit of sheep α2M was found to be made up of two fragments of 102 and 31 kDa respectively. The proteinase inhibitor from sheep was found to have Stokes radius of 79Ǻ, which makes it much more compact than its human homologue. It entraps only 1 mol of trypsin per mole of inhibitor, like its caprine counterpart. The use of isothermal titration calorimetry has become gold standard for exploring thermodynamics of binding interactions. In this study, binding interaction of trypsin with alpha-2-macroglobulin is studied using ITC. The thermodynamic signatures--enthalpy change (ΔH), entropy change (ΔS) and Gibb's free energy change (ΔG), along with number of binding sites (N) and affinity constant (K) are explored for α2M-trypsin binding for the first time for any known α2M molecule. The thermodynamics of proteinase-antiproteinase association suggests that trypsin-α2M interaction is enthalpy driven event., (Copyright © 2015 Elsevier B.V. All rights reserved.)