20S HUMAN PROTEASOMES BIND WITH A SPECIFIC ORIENTATION TO LIPID MONOLAYERS IN-VITRO

20S Proteasomes are non-lysosomal, high molecular weight proteinases i mplicated in the degradation of misfolded proteins and several short-l ived regulatory proteins. They have a well established role, as the co re of the 26S proteasome complex, in the ubiquitin-dependent proteolyt ic pathway and in antigen processing. While correctly folded proteins are not degraded by the 20S proteasome, unfolding, for example by oxid ation, may render them degradable. The 20S proteasome is a 700-kDa cyl indrical particle, composed of 14 subunits of molecular masses 20-35 k Da. There is evidence that 20S proteasomes are in close proximity to o r associate with the endoplasmic reticulum and nuclear and plasma memb ranes in vivo. To better understand the lipid association of 20S prote asomes in vitro, we used a lipid monolayer system as a simple model sy stem for biological membranes. The structure and orientation of the mo nolayer lipid bound 20S proteasomes has been determined by electron mi croscopy. 20S proteasomes associated in an 'end-on' configuration spec ifically on PI lipid monolayers forming large arrays, with their chann els opposite the lipid headgroups. On ER and Golgi lipid films 20S pro teasomes were oriented in the same way as on the PI lipid film but wer e monodisperse. Protein molecules were randomly oriented in the presen ce of PA, PG, PS, PC and mitochondrial lipid monolayers. We show that 20S proteasomes bind to phospholipids in vitro in a preferred orientat ion which places the proteasome channel perpendicular to the membrane.