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HspA1A, a seventy‐kDa heat shock protein, binds to phosphoinositides
Author(s) -
Kdeiss Brianna,
McCallister Chelsea,
Nikolaidis Nikolas
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.661.3
Subject(s) - phosphatidylserine , biochemistry , plasma protein binding , heat shock protein , biology , hsp70 , microbiology and biotechnology , binding site , chemistry , membrane , phospholipid , gene
Seventy‐kilodalton heat shock proteins (Hsp70s) are molecular chaperones essential for maintaining cellular homeostasis. Apart from their indispensable functions in protein folding and refolding Hsp70s also localize at the plasma membrane and bind to specific lipids. The interaction of Hsp70s with lipids has direct physiological outcomes including lysosomal rescue, microautophagy, and promotion of cell apoptosis. We have recently characterized the binding properties of HspA1A, a major inducible Hsp70 in humans and mice, to several lipids, including phosphatidylserine and sulfatide. However, whether HspA1A binds to lipids essential for cell signaling, like phosphoinositides, remains unknown. Therefore, in this study, we characterized the interaction of HspA1A with several such lipids. We first established that HspA1A binds to monophosphorylated inositides with much higher affinity than to di‐ and triphosphorylated inositides. We also used high concentrations of potassium and found that HspA1A embeds in membranes when bound to specific lipids. We also determined that the nucleotide‐binding domain (NBD) of the protein binds to lipids quantitatively more than the substrate‐binding domain (SBD). However, for all lipids studied, the full‐length protein is necessary for binding. Furthermore, we used calcium and reaction buffers equilibrated at different pH values and determined that electrostatic interactions alone cannot fully explain the association of HspA1A with phosphoinositides. We then determined that lipid‐binding is inhibited by nucleotide‐binding, but it is not significantly reduced by protein‐substrate binding. Together, these results suggest that the interaction of HspA1A with phosphoinositides is specific, depends on the physicochemical properties of the lipid, and is mediated by multiple molecular forces and binding sites. Based on these data we propose a model according to which the NBD domain of HspA1A contains two lipid binding sites, the first one targets and docks the protein to the membrane, while the second one facilitates the embedding of the SBD into the membrane. These results uncover mechanistic details of the Hsp70‐lipid interactions and establish a framework of possible physiological functions of these interactions as they relate to chaperone mediated cell signaling and communication.