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Liquid-Liquid Phase Transition of Protein Aqueous Solutions Isothermally Induced by Protein Cross-Linking
 

Summary: Liquid-Liquid Phase Transition of Protein Aqueous Solutions
Isothermally Induced by Protein Cross-Linking
Ying Wang and Onofrio Annunziata*
Department of Chemistry, Texas Christian UniVersity, Fort Worth, Texas 76129
ReceiVed October 16, 2007. In Final Form: NoVember 28, 2007
We experimentally demonstrated that liquid-liquid phase separation (LLPS) of protein aqueous solutions can be
induced by isothermal protein oligomerization. This phenomenon is analogous to LLPS induced by the polymerization
of small organic molecules in solution. Specifically, using glutaraldehyde for protein cross-linking, we observed the
formation of protein-rich liquid droplets for bovine serum albumin and chicken egg lysozyme at 25 C. These droplets
evolved into cross-linked protein microspheres. If the aqueous solutions of the protein monomer do not show LLPS
at temperatures lower than the oligomerization temperature, protein-rich droplets are not observed. We experimentally
linked the formation of these droplets to the increase of LLPS temperature during protein oligomerization. When
macroscopic aggregation competes with LLPS, a rationale choice of pH, polyethylene glycol, and salt concentrations
can be used to favor LLPS relative to aggregation. Although glutaraldehyde has been extensively used to cross-link
protein molecules, to our knowledge, its use in homogeneous aqueous solutions to induce LLPS has not been previously
described. This work contributes to the fundamental understanding of both phase transitions of protein solutions and
the morphology of protein condensed phases. It also provides guidance for the development of new methods based
on mild experimental conditions for the preparation of protein-based materials.
Introduction
Isothermalpolymerizationreactionsofsmallorganicmolecules

  

Source: Annunziata, Onofrio - Department of Chemistry, Texas Christian University
Benedek, George B. - Department of Physics, Massachusetts Institute of Technology (MIT)

 

Collections: Chemistry; Physics