Human and Bacterium Hsp70 Interacts Differently with Lipid Membranes

Abstract

The cellular response to stress is orchestrated by the expression of a family of proteins coined heat shock proteins (hsp), which are involved in the stabilization of basic cellular processes to preserve cell viability and homeostasis. The bulk of hsp function occurred within the cytosol and subcellular compartments. However, some hsp have also been found outside cells released by an active mechanism independent of cell death. Extracellular hsp act as signaling molecules directed at activating a systemic response to stress. The export of hsp requires the translocation from the cytosol into the extracellular milieu across the plasma membrane. We have proposed that membrane insertion is the initial step in the export process. We investigated the interaction of the major inducible hsp, Hsp70, from humans (HspA1) and bacteria (DnaK) with liposomes. We found that HspA1 displayed a high specificity for negatively charged phospholipids, such as phosphatidyl serine, whereas DnaK interacted with all lipids tested regardless of the charge. Both proteins were inserted into the lipid bilayer as demonstrated by resistance to acid or basic washes, which was confirmed by partial protection from proteolytic cleavage. The majority of HspA1 was inserted into the membrane with a small region of the N-terminus end exposed to the outer phase of the liposome. In contrast, the N-terminus end of DnaK was inserted into the membrane, exposing the C-terminus end outside the liposome. HspA1 was found to make high oligomeric complexes upon insertion into the membranes whereas DnaK only formed dimers within the lipid membrane. These observations suggest that both Hsp70s interact with lipids, but mammalian HspA1 displayed a high degree of specificity and structure as compared with the bacterium form.