Information on EC 3.6.3.51 - mitochondrial protein-transporting ATPase and Organism(s) Homo sapiens

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Homo sapiens


The expected taxonomic range for this enzyme is: Eukaryota, Bacteria


The taxonomic range for the selected organisms is: Homo sapiens

EC NUMBER
COMMENTARY hide
3.6.3.51
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RECOMMENDED NAME
GeneOntology No.
mitochondrial protein-transporting ATPase
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of phosphoric ester
transmembrane transport
SYSTEMATIC NAME
IUBMB Comments
ATP phosphohydrolase (mitochondrial protein-importing)
A non-phosphorylated, non-ABC (ATP-binding cassette) ATPase involved in the transport of proteins or preproteins into mitochondria using the TIM protein complex. (TIM is the protein transport machinery of the inner mitochondrial membrane that contains three essential Tim proteins: Tim17 and Tim23 are thought to build a preprotein translocation channel while Tim44 interacts transiently with the matrix heat-shock protein Hsp70 to form an ATP-driven import motor.)
CAS REGISTRY NUMBER
COMMENTARY hide
9000-83-3
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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the presequence-cleaved mature forms of the recombinant fusion proteins with a 50 or 60 residue spacer sequence are proteinase K-resistant, whereas those with a 70 or 80 residue spacer sequence are proteinase K-sensitive
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
GrpEL1
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the proposed nucleotide exchange factor of the human mtHsp70 machine. To access the nucleotide relase of human GrpEL1, a single turnover ATPase assay is performed to monitor the hydrolysis of the prebound mtHsp70-ATP complex of the wild-type protein in the presence of GrpEL1. Human GrpEL1 shows robust nucleotide exchange activity in the presence of excess unlabeled ATP, thus inhibiting the maximum ATP hydrolysis of wild-type mtHsp70
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Hsp40-related J domain protein DJA1
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Hsp40-related J domain protein DJA2
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Hsp40-related J domain protein DJA4
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human escort protein
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Hep, stimulates the activity of mtHsp70 49fold, and also 11.5fold the activity of the isolated ATPase domain. Hep binding to full-length mtHsp70 and its isolated ATPase domain is strongest in the absence of nucleotides
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human Hsp70 escort protein
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human Hsp70 escort protein (Hep) possesses the unique ability to stimulate the ATPase activity of mtHsp70 as well as to prevent the aggregation of unfolded client proteins similar to J-proteins
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J-protein
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J-protein splice hTid-1L, larger isoform and hTid-1S, smaller isoform. At a morlar ratio of mtHsp70 to J-protein, a 3.2fold stimulation is observed with hTid-1L for wild-type protein. hTid-1S shows an 11fold stimulation for wild-type mtHp70
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TIM14
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stimulates
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00068
ATP
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pH 7.5, 25°C, full-length mtHsp70 in absence of Hep
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30
ATPase activity assay
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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TIG-1 cell, overexpression of mortalin extends the in vitro lifespan of the fibroblasts
Manually annotated by BRENDA team
additional information
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NARF cell, stably transfected with isopropyl beta-D-thiogalactoside-inducible p14ARF
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
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matrix
Manually annotated by BRENDA team
PDB
SCOP
CATH
UNIPROT
ORGANISM
Homo sapiens;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
81000
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1 * 81000, full-length enzyme in absence of ADP
86000
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1 * 86000, full-legth enzyme in presence of ADP, SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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full-legth enzyme in presence of ADP, SDS-PAGE
monomer
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1 * 81000, full-length enzyme in absence of ADP; 1 * 86000, full-legth enzyme in presence of ADP, SDS-PAGE
additional information
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in absence of nucleotides, Hsp70 also forms oligomeric aggregates of high molecular weight
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
mtHsp70 and Hsp70 ATPase domain remain soluble in the absence of Hep and nucleotides when stored in Tris at pH 8.0. In the case of the ATPase domain, the protein remains soluble at concentrations as high as 0.2 mM even when stored for 24 h at room temperature. However, the mtHsp70 ATPase domain could not be stably stored at high concentrations for a similar period of time in buffers having pH values more closely resembling the physiological environment within Escherichia coli where these proteins are overexpressed, unless it is incubated with equimolar Hep
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified on nickel-Sepharose, by ion exchange on a Mono Q column, and by gel filtration
recombinant His-tagged full-length enzyme and isolated ATPase domain from Escherichia coli by nickel affinity chromatography, ammonium sulfate fractionation, anion exchange chromatography, and gel filtration
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recombinant His-tagged mtHsp70
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression of mtHsp70 and its isolated ATPase domain, as well as the ATPase fused to the peptide-binding domain of HscA, in Escherichia coli as insoluble proteins; HEK-293 cells are transiently transfected with pHep-EGFP, expression of C-terminally His-tagged Hsp70 without its N-terminal mitochondrial targeting sequence in Escherichia coli, the enzyme is insoluble in absence of human escort protein Hep, also as isolated ATPase domain and a chimera having this domain fused to the peptide-binding domain of HscA, a soluble monomeric chaperone
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for purification of the His-tagged human mtHsp70, coexpression is carried out with yeast Hsp70 escort protein Hep in Escherichia coli BL21(DE3)
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the sequence encoding human Hsc70 is inserted into the pProExHTa vector
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Y196A/N198A/D199A
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YND mutant, shows 3fold elevated basal activity in comparison with wild-type. The mutant also shows a significant decrease in Hsp70 escort protein Hep stimulation as in comparison to the wild-type protein. The YND mutant also exhibits a decreased stimulation of the J-protein splice variant hTid-1S in comparison of the wild-type further supporting the mutually exclusive nature of Hep and J-protein interaction at the ATPase domain of human mtHsp70
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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in vivo binding of mortalin/mtHsp70 with HSP60, involvment in tumorigenesis, functional distinction in pathways involved in senescence
additional information
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TIM44 binds to the peptide-binding domain of Hsp70 and thereby recruits it to the outlet of the translocation pore, so that mtHsp70 can bind to the incoming polypeptide chain as soon as it emerges from the channel