Effects of pH on protein-protein interactions and implications for protein phase behavior.
The effects of pH on protein interactions and protein phase behavior were investigated by measuring the reduced second osmotic virial coefficient (b2) for ovalbumin and catalase, and the aggregate and crystal solubilities for ovalbumin, beta-lactoglobulin A and B, ribonuclease A and lysozyme. The b2 trends observed for ovalbumin and catalase show that protein interactions become increasingly attractive with decreasing pH. This trend is in good agreement with ovalbumin phase behavior, which was observed to evolve progressively with decreasing pH, leading to formation of amorphous aggregates instead of gel bead-like aggregates, and spherulites instead of needle-like crystals. For both acidic and basic proteins, the aggregate solubility during protein salting-out decreased with decreasing pH, and contrary to what is commonly believed, neither aggregate nor crystal solubility had a minimum at the isoelectric point. beta-Lactoglobulin B was the only protein investigated to show salting-in behavior, and crystals were obtained at low salt concentrations in the vicinity of its isoelectric point. The physical origin of the different trends observed during protein salting-in and salting-out is discussed, and the implications for protein crystallization are emphasized.
Dumetz
André C
AC
Center for Molecular and Engineering Thermodynamics, Department of Chemical Engineering, University of Delaware, Newark, DE 19716, USA.
Chockla
Aaron M
AM
Kaler
Eric W
EW
Lenhoff
Abraham M
AM
D000465:Algorithms; D000818:Animals; D002374:Catalase / Q000737:chemistry / Q000378:metabolism; D002417:Cattle; D002645:Chickens; D002845:Chromatography / Q000379:methods; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007782:Lactoglobulins / Q000737:chemistry / Q000378:metabolism; D009113:Muramidase / Q000737:chemistry / Q000378:metabolism; D010047:Ovalbumin / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000737:chemistry / Q000378:metabolism; D012995:Solubility
D007782:Lactoglobulins; D011506:Proteins; D010047:Ovalbumin; D002374:Catalase; D012259:Ribonuclease, Pancreatic; D009113:Muramidase