Hydrostatic pressure rescues native protein from aggregates

Misfolding and misassembly of proteins are major problems in the biotechnol ogy industry, in biochemical research, and in human disease. Here we descri be a novel approach for reversing aggregation and increasing refolding by a pplication of hydrostatic pressure. Using P22 tailspike protein as a model system, intermediates along the aggregation pathway were identified and qua ntitated by size-exclusion high-performance liquid chromatography (HPLC). T ailspike aggregates were subjected to hydrostatic pressures of 2.4 kbar (35 ,000 psi). This treatment dissociated the tailspike aggregates and resulted in increased formation of native trimers once pressure was released. Tails pike trimers refolded at these pressures were fully active for formation of infectious viral particles. This technique can facilitate conversion of ag gregates to native proteins without addition of chaotropic agents, changes in buffer, or large-scale dilution of reagents required for traditional ref olding methods. Our results also indicate that one or more intermediates at the junction between the folding and aggregation pathways is pressure sens itive. This finding supports the hypothesis that specific determinants of r ecognition exist for protein aggregation, and that these determinants are s imilar to those involved in folding to the native state. An increased under standing of this specificity should lead to improved refolding methods. (C) 1999 John Wiley & Sons, Inc.