title: Refolding of urea denatured ovalbumin with three phase partitioning generates many conformational variants.
abstract: Three phase partitioning is a process in which mixing t-butanol with ammonium sulphate with a protein solution leads to the formation of three phases. Generally, the interfacial protein precipitate (formed between upper t-butanol rich and lower aqueous phase) can be easily dissolved back in aqueous buffers. In case of ovalbumin, this led to a precipitate which was insoluble in aqueous buffers. This precipitate when solubilized with 8 M urea and subjected to three phase partitioning under various conditions led to many refolded soluble conformational variants of ovalbumin. One of these showed trypsin inhibitory activity, had marginally higher β-sheet content and had higher surface hydrophobicity (both with respect to native ovalbumin). Scanning electron microscopy and Atomic force microscopy of this preparation showed a thread like structure characteristic of amyloid fibrils. The behaviour of ovalbumin during three phase partitioning makes it a valuable system for gaining further understanding of protein aggregation.
authors.affiliation: Chemistry Department, Indian Institute of Technology Delhi, Hauz Khas, New Delhi 110 016, India.
authors.forename: Gulam Mohmad
authors.initials: GM
authors.lastname: Rather
authors.forename: Munishwar Nath
authors.initials: MN
authors.lastname: Gupta
mesh_terms: D000818:Animals; D002645:Chickens; D004529:Egg White; D057927:Hydrophobic and Hydrophilic Interactions; D010047:Ovalbumin / Q000737:chemistry* / Q000648:ultrastructure; D011487:Protein Conformation; D011489:Protein Denaturation; D058767:Protein Unfolding; D013816:Thermodynamics; D014508:Urea / Q000737:chemistry*
chemical_list: D014508:Urea; D010047:Ovalbumin
keywords: Amyloid fibrils; Ovalbumin; Protein aggregation; Protein refolding; Three phase partitioning