Correlation for the partition behavior of proteins in aqueous two‐phase systems: Effect of surface hydrophobicity and charge

Abstract

Correlations to describe the effect of surface hydrophobicity and charge of proteins with their partition coefficient in aqueous two-phase systems were investigated. Polyethylene glycol (PEG) 4000/phosphate, sulfate, citrate, and dextran systems in the presence of low (0.6% w/w) and high (8.8% w/w) levels of NaCl were selected for a systematic study of 12 proteins. The surface hydrophobicity of the proteins was measured by ammonium sulfate precipitation as the inverse of their solubility. The hydrophobicity values measured correlated well with the partition coefficients, K, obtained in the PEG/salt systems at high concentration of NaCl (r = 0.92-0.93). In PEG/citrate systems the partition coefficient correlated well with protein hydrophobicity at low and high concentrations of NaCl (r = 0.81 and 0.93, respectively). The PEG/citrate system also had a higher hydrophobic resolution than other systems to exploit differences in the protein's hydrophobicity. The surface charge and charge density of the proteins was determined over a range of pH (3-9) by electrophoretic titration curves; PEG/salt systems did not discriminate well between proteins of different charge or charge density. In the absence of NaCl, K decreased slightly with increased positive charge. At high NaCl concentration, K increased as a function of positive charge. This suggested that the PEG-rich top phase became more negative as the concentration of NaCl in the systems increased and, therefore, attracted the positively charged proteins. The effect of charge was more important in PEG/dextran systems at low concentrations of NaCl. In the PEG/dextran systems at lower concentration of NaCl, molecular weight appeared to be the prime determinant of partition, whereas no clear effect of molecular weight could be found in PEG/salt systems.

Extraction and Purification of Bioproducts and Nanoparticles using Aqueous Two‐Phase Systems Strategies

“…Although research has been conducted in this area to facilitate process design and reduce the requirement for time-consuming experiments, the true molecular understanding of solute partitioning is just beginning to be developed. The establishment of guidelines has allowed the prediction of the partition behavior of solutes in ATPS based on their physicochemical properties [1,5,6]. Three physicochemical properties have been identified to be mainly responsible for governing solute partition behavior in ATPS: (1) molecular weight, (2) hydrophobicity, and (3) superficial net electrochemical charge [1,5].…”

“…The establishment of guidelines has allowed the prediction of the partition behavior of solutes in ATPS based on their physicochemical properties [1,5,6]. Three physicochemical properties have been identified to be mainly responsible for governing solute partition behavior in ATPS: (1) molecular weight, (2) hydrophobicity, and (3) superficial net electrochemical charge [1,5]. It is clear that once solute partitioning has been characterized, a process can be designed for the primary recovery and partial purification of the product of interest.…”

Isolation and identification of a high molecular weight protein in sow milk

“…Therefore, HMWP should contain all 20 amino acids. By SDS-PAGE, gel filtration chromatography, pI determination, amino acid assay and references (Kontopidis et al, 2004;Andrews et al, 2005), HMWP appears to be a soluble, acidic and monomeric globulin. However, in order to more fully understand the biochemical properties and biological function of HMWP, further study is required.…”

Characterization of Feed Properties for Conceptual Process Design Involving Complex Mixtures, Such as Natural Extracts

“…3) Experimental determination Equilibrium phase distribution coefficients and separation factors for the individual unit operation are determined in laboratory-scale experiments [21,24,[32][33][34][35][36][37][38][39].…”