Human heat shock protein 70 (Hsp70) as a peripheral membrane protein |
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| Authors: | Ajay K. Mahalka,Thomas Kirkegaard,Laura T.I. Jukola,Marja Jä ä ttelä ,Paavo K.J. Kinnunen |
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| Affiliation: | 1. Helsinki Biophysics and Biomembrane Group, Department of Biomedical Engineering and Computational Science, Aalto University, Espoo, Finland;2. Cell Death and Metabolism, Danish Cancer Society Research Center, Copenhagen, Denmark;3. Orphazyme ApS, Copenhagen, Denmark |
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| Abstract: | While a significant fraction of heat shock protein 70 (Hsp70) is membrane associated in lysosomes, mitochondria, and the outer surface of cancer cells, the mechanisms of interaction have remained elusive, with no conclusive demonstration of a protein receptor. Hsp70 contains two Trps, W90 and W580, in its N-terminal nucleotide binding domain (NBD), and the C-terminal substrate binding domain (SBD), respectively. Our fluorescence spectroscopy study using Hsp70 and its W90F and W580F mutants, and Hsp70-?SBD and Hsp70-?NBD constructs, revealed that binding to liposomes depends on their lipid composition and involves both NBD and SBD. |
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| Keywords: | AcrA, acrylamide aSM, acid sphingomyelinase a.u., arbitrary unit BMP, bis(monoacylglycero)phosphate Br2PC, brominated phosphatidylcholine 6,7Br2-PC, 1-palmitoyl-2-(6,7-dibromo)stearoyl-sn-glycero-3-phosphocholine 9,10Br2-PC, 1-palmitoyl-2-(9,10-dibromo)stearoyl-sn-glycero-3-phosphocholine 11,12Br2-PC, 1-palmitoyl-2-(11,12-dibromo)stearoyl-sn-glycero-3-phosphocholine Br4BMP, bis[mono(9,10)-dibromostearoyl]glycerophosphate 9,10Br2-PS, 1-palmitoyl-2-(9,10-dibromo)stearoyl-sn-glycero-3-phospho-l-serine Br8CL, tetra(9,10-dibromo stearoyl)cardiolipin Br2PS, brominated phosphatidylserine CD, circular dichroism Chol, cholesterol CL, cardiolipin DnaK, E. coli heat shock protein 70 DTT, dithiothreitol EDTA, ethylenediamine-N,N,N&prime ,N&prime -tetraacetic acid F, fluorescence intensity F0, initial fluorescence intensity FA, fatty acid Grp78, endoplasmic reticulum heat shock protein 70 HD, Huntington disease HDP, host defense peptides Hepes, 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid Hsc70, constitutively expressed heat shock protein 70 Hsp70, Heat shock protein of &asymp 70 kDa Hsp70-?NBD, recombinant Hsp70 lacking the nucleotide binding domain Hsp70-?SBD, recombinant Hsp70 lacking the substrate binding domain Hsp70-W90F, recombinant Hsp70 with substitution W90F Hsp70-W580F, recombinant Hsp70 with substitution W580F KCL, potassium chloride Ksv, Stern&ndash Volmer quenching constants L/P, lipid/protein molar ratio LUV, large unilamellar vesicles MES, 2-(N-morpholino)ethanesulfonic acid NBD, nucleotide binding domain NPD, Niemann&ndash Pick disease PEG, polyethylene glycol POPC, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine POPS, 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine PS, phosphatidylserine RFI, relative fluorescence intensity Spm, sphingomyelin SBD, substrate binding domain Ssa1p, S. cerevisiae heat shock protein 70 tOCL, 1,1&prime ,2,2&prime -tetraoleoyl cardiolipin wtHsp70, wild type Hsp70 π, surface pressure π0, initial surface pressure ?π, increment in surface pressure πc, critical packing pressure λ, wavelength ?λ, spectral center of mass |
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