Structural and Biological Interaction of hsc-70 Protein with Phosphatidylserine in Endosomal Microautophagy.

The Journal of biological chemistry 2016 Aug 26

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hsc-70 (HSPA8) is a cytosolic molecular chaperone, which plays a central role in cellular proteostasis, including quality control during protein refolding and regulation of protein degradation. hsc-70 is pivotal to the process of macroautophagy, chaperone-mediated autophagy, and endosomal microautophagy. The latter requires hsc-70 interaction with negatively charged phosphatidylserine (PS) at the endosomal limiting membrane. Herein, by combining plasmon resonance, NMR spectroscopy, and amino acid mutagenesis, we mapped the C terminus of the hsc-70 LID domain as the structural interface interacting with endosomal PS, and we estimated an hsc-70/PS equilibrium dissociation constant of 4.7 ± 0.1 μm. This interaction is specific and involves a total of 4-5 lysine residues. Plasmon resonance and NMR results were further experimentally validated by hsc-70 endosomal binding experiments and endosomal microautophagy assays. The discovery of this previously unknown contact surface for hsc-70 in this work elucidates the mechanism of hsc-70 PS/membrane interaction for cytosolic cargo internalization into endosomes. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Citation

Kateryna Morozova, Cristina C Clement, Susmita Kaushik, Barbara Stiller, Esperanza Arias, Atta Ahmad, Jennifer N Rauch, Victor Chatterjee, Chiara Melis, Brian Scharf, Jason E Gestwicki, Ana-Maria Cuervo, Erik R P Zuiderweg, Laura Santambrogio. Structural and Biological Interaction of hsc-70 Protein with Phosphatidylserine in Endosomal Microautophagy. The Journal of biological chemistry. 2016 Aug 26;291(35):18096-106