Pressure-Induced Critical Association of Myoglobin R. Gebhardt, W. Doster, J. Friedrich*, W. Petry and A. Schulte Technical University Munich, Physics Department E13 and E 14*, D-85748 Garching, Germany, wdoster@ph.tum.de , presented at the High Pressure Bioscience and Biotechnol- ogy Conference 2002 in Dortmund Abstract Oligomeric proteins generally tend to dissociate under pressure, while aggregation is often observed in the pressure-denatured state. We report on a pressure-induced association of native myoglobin in the low-pressure regime. Depending on solvent conditions, the monomer becomes unstable above a critical pressure. The associa- tion process involves an initial lag phase due to seed formation, an intermediate re- versible oligomeric state and a final structurally altered precipitate. Using static and dynamic light scattering, we discuss the kinetics of association, the effect of pH, temperature and buffer conditions on the stability limit. Introduction Aggregation processes are of increasing interest because of their relevance to a number of human deseases e.g. Alzheimers's disease or bovine spongiform en- cephalopathy (BSE). The relevant proteins take up an incorrect, fibrillar or amour- phous structure. Besides misfolding, chemical denaturants, temperature or pres- sure-denaturation lead to so-called folding-intermediates with the tendency to aggregate [1,2]. One important goal in attempts to understand the events occurring during the amyloid formation is to find out the critical conditions were aggregation occurs and to uncover possibilities to modulate their interactions. In the present work we explore the pressure-dependence of the met-myoglobin aggregation proc- ess over a wide pH-range. The pressure-denaturation of sperm-whale met- myoglobin, and its relation to precipitation was investigated by Zipp and Kauzmann [3]. Aggregation was never observed below pH 5 or above pH 9. Re- cently, the formation of myoglobin-aggregates after pressure-treatment has been reported [4,5]. High pressure leads to an intermediate, which tends to aggregate if the pressure is reduced to ambient values. We report however, that myoglobin can associate under native conditions in the low-pressure regime within a narrow pH range. The initial phase of the association is reversible, but irreversible aggregates are formed which exhibit intermolecular β-sheets at the final stage.