Abstract
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.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 18096-18106 |
| Number of pages | 11 |
| Journal | Journal of Biological Chemistry |
| Volume | 291 |
| Issue number | 35 |
| DOIs | |
| State | Published - Aug 26 2016 |
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ASJC Scopus subject areas
- Biochemistry
- Medicine(all)
- Molecular Biology
- Cell Biology
Cite this
Structural and biological interaction of HSC-70 protein with phosphatidylserine in endosomal microautophagy. / Morozova, Kateryna; Clement, Cristina C.; Kaushik, Susmita; Stiller, Barbara; Arias, Esperanza; Ahmad, Atta; Rauch, Jennifer N.; Chatterjee, Victor; Melis, Chiara; Scharf, Brian; Gestwicki, Jason E.; Cuervo, Ana Maria; Zuiderweg, Erik R P; Santambrogio, Laura.
In: Journal of Biological Chemistry, Vol. 291, No. 35, 26.08.2016, p. 18096-18106.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Structural and biological interaction of HSC-70 protein with phosphatidylserine in endosomal microautophagy
AU - Morozova, Kateryna
AU - Clement, Cristina C.
AU - Kaushik, Susmita
AU - Stiller, Barbara
AU - Arias, Esperanza
AU - Ahmad, Atta
AU - Rauch, Jennifer N.
AU - Chatterjee, Victor
AU - Melis, Chiara
AU - Scharf, Brian
AU - Gestwicki, Jason E.
AU - Cuervo, Ana Maria
AU - Zuiderweg, Erik R P
AU - Santambrogio, Laura
PY - 2016/8/26
Y1 - 2016/8/26
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=84984783742&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84984783742&partnerID=8YFLogxK
U2 - 10.1074/jbc.M116.736744
DO - 10.1074/jbc.M116.736744
M3 - Article
C2 - 27405763
AN - SCOPUS:84984783742
VL - 291
SP - 18096
EP - 18106
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 35
ER -