title: Identification of a new alpha-2-macroglobulin: Multi-spectroscopic and isothermal titration calorimetry study.
abstract: 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.
authors.affiliation: Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh 202002, India.
authors.forename: Ahmed Abdur
authors.initials: AA
authors.lastname: Rehman
authors.affiliation: Department of Biochemistry, Faculty of Dentistry, Jamia Millia Islamia, New Delhi 110025, India.
authors.forename: Haseeb
authors.initials: H
authors.lastname: Ahsan
authors.affiliation: Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh 202002, India. Electronic address: fahimhkhan@rediffmail.com.
authors.forename: Fahim Halim
authors.initials: FH
authors.lastname: Khan
mesh_terms: D000818:Animals; D002151:Calorimetry; D002241:Carbohydrates / Q000032:analysis; D019277:Entropy; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008744:Methylamines / Q000494:pharmacology; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D011487:Protein Conformation; D012756:Sheep; D013057:Spectrum Analysis; D014357:Trypsin / Q000378:metabolism; D000511:alpha-Macroglobulins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D002241:Carbohydrates; D008744:Methylamines; D011480:Protease Inhibitors; D000511:alpha-Macroglobulins; C027451:methylamine; D014357:Trypsin
keywords: Alpha macroglobulin; Alpha-2-macroglobulin; Antiproteinase; Biochemical characterisation; ITC; Ovis aries