Pressure Effect on the Temperature-Induced Unfolding and Tendency To Aggregate of Myoglobin

Abstract

This work demonstrates that pressure-induced partially unfolded states play a very important role in the aggregation of proteins. The high-pressure unfolding of horse heart metmyoglobin results in an intermediate form that shows a strong tendency to aggregate after pressure release. These aggregates are similar to those that are usually observed upon temperature denaturation. Infrared spectra in the amide I region indicate the formation of an intermolecular antiparallel beta-sheet stabilized by hydrogen bonding. The formation of the aggregates is temperature-dependent. Below 30 degrees C, no aggregation is taking place as seen from the infrared spectra. At 45 and 60 degrees C, two types of aggregates are formed: one that can be dissociated by moderate pressures and one that is pressure-insensitive. When precompressed at 5 degrees C, temperature-induced aggregation takes place at lower temperature (38 degrees C) than without pressure pretreatment (74 degrees C).

High pressure fosters protein refolding from aggregates at high concentrations

“…In fact, recovery of both rhGH and lysozyme exhibited independence of protein concentration in the ranges studied. This finding is consistent with previous reports that the pressure-induced dissociation of erythrocruorin is concentration independent (43), as is the dissociation at 3.5 kbar of myoglobin aggregates initially formed by unfolding at 12 kbar (44). Previous refolding studies at atmospheric conditions report strong negative dependence of recovery of native protein on protein concentration, even when concentrations were very low (e.g., usually 1-50 g͞ml) (14,33).…”

Protein dynamics: hydration and cavities

“…Further evidence for the absence of water in the fibrils comes from unpublished hydrogen-deuterium exchange experiments on insulin fibrils that show that the exchange is slow even at high pressures. Interestingly, a similar difference in the pressure stability of early and mature aggregates has been observed for the amorphous aggregates of myoglobin (57) and an amyloid-like structure of the prion protein obtained by thermal treatment (53).…”

The preaggregated state of an amyloidogenic protein: Hydrostatic pressure converts native transthyretin into the amyloidogenic state

“…This finding is important, because it opens the possibility of carrying out structural studies (such as highresolution NMR) after pressure treatment at 1°C to characterize this amyloidogenic intermediate. Recently, Smeller et al (38) has shown that myoglobin, a nonamyloidogenic protein, also undergoes aggregation after pressure denaturation at 12 kbar.…”