Characterisation of several Hsp70 interacting proteins from mammalian organelles.
Since both the spectrum and characteristics of in vivo substrates with affinity for Hsp70 members are largely unknown, we have investigated the range and type of mammalian organellar proteins which selectively interact with immobilised Escherichia coli Hsp70 (DnaK). Amongst a subset of organellar proteins selectively retained on DnaK, the major constituents represent unstable proteins and subunits of oligomeric proteins. The interactions with DnaK were diminished in the presence of mt-Hsp70 and BiP, while the complexes formed with DnaK were dissociated in the presence of K+ and GrpE-like co-chaperones, suggesting that these organellar proteins constitute general Hsp70 substrates. Protein sequence analysis identified the major DnaK interacting constituents as the mitochondrial transcription factor A, the alpha- (but not the beta-) subunit of succinyl CoA synthetase, mitochondrial 2,4-dienoyl CoA reductase, endoplasmic reticulum cyclophilin-B, peroxisomal multifunctional enzyme and a previously undescribed peroxisomal protein suspected to represent an isoform of 2,4-dienoyl CoA reductase. The selective retention of these fully synthesised proteins on Hsp70 most likely reflects the function of this molecular chaperone in protein biogenesis, but additionally, could extend the known functions of Hsp70 to include modulating the activities of certain proteins or enzymes which are important in cellular homeostasis.
Naylor
D J
DJ
Department of Horticulture, Viticulture and Oenology, The University of Adelaide, Waite campus, PMB1, Glen Osmond, SA 5064, Australia. dnaylor@waite.adelaide.edu.au
Hoogenraad
N J
NJ
Høj
P B
PB
D000255:Adenosine Triphosphate; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry*; D006360:Heat-Shock Proteins / Q000302:isolation & purification; D008099:Liver / Q000737:chemistry; D008928:Mitochondria / Q000737:chemistry; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D015388:Organelles / Q000737:chemistry*; D011189:Potassium Chloride; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013552:Swine
D029968:Escherichia coli Proteins; C109172:GrpE-like 1 mitochondrial protein, vertebrate; C484655:Grpel1 protein, rat; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D011506:Proteins; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; C063842:dnaK protein, E coli