Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P11021 (BiP)
 2,049 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The huntingtin exon 1 proteins with a polyglutamine repeat in the pathological range (51 or 83 glutamines), but not with a polyglutamine tract in the normal range (20 glutamines), form aggresome-like perinuclear inclusions in human 293 Tet-Off cells. These structures contain aggregated, ubiquitinated huntingtin exon 1 protein with a characteristic fibrillar morphology. Inclusion bodies with truncated huntingtin protein are formed at centrosomes and are surrounded by vimentin filaments. Inhibition of proteasome activity resulted in a twofold increase in the amount of ubiquitinated, SDS-resistant aggregates, indicating that inclusion bodies accumulate when the capacity of the ubiquitin-proteasome system to degrade aggregation-prone huntingtin protein is exhausted. Immunofluorescence and electron microscopy with immunogold labeling revealed that the 20S, 19S, and 11S subunits of the 26S proteasome, the molecular chaperones BiP/GRP78, Hsp70, and Hsp40, as well as the RNA-binding protein TIA-1, the potential chaperone 14-3-3, and alpha-synuclein colocalize with the perinuclear inclusions. In 293 Tet-Off cells, inclusion body formation also resulted in cell toxicity and dramatic ultrastructural changes such as indentations and disruption of the nuclear envelope. Concentration of mitochondria around the inclusions and cytoplasmic vacuolation were also observed. Together these findings support the hypothesis that the ATP-dependent ubiquitin-proteasome system is a potential target for therapeutic interventions in glutamine repeat disorders.
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PMID:Accumulation of mutant huntingtin fragments in aggresome-like inclusion bodies as a result of insufficient protein degradation. 1135 30

Intracellular Ca(2+) in Trypanosoma cruzi is mainly located in an acidic compartment named the acidocalcisome, which among other pumps and exchangers possesses a plasma membrane-type Ca(2+)-ATPase. Evidence for an endoplasmic reticulum-located Ca(2+) uptake has been more elusive and based on indirect results. Here we report the cloning and sequencing of a gene encoding a sarcoplasmic-endoplasmic reticulum-type Ca(2+)-ATPase from T. cruzi. The protein (TcSCA) predicted from the nucleotide sequence of the gene has 1006 amino acids and a molecular mass of 109.7 kDa. Several sequence motifs found in sarcoplasmic-endoplasmic reticulum-type Ca(2+)-ATPases were present in TcSCA. Expression of TcSCA in yeast mutants deficient in the Golgi and vacuolar Ca(2+) pumps (pmr1 pmc1 cnb 1) restored growth on EGTA. Membranes were isolated from the pmr1 pmc1 cnb1 mutant transformed with TcSCA, and it was found that the TcSCA polypeptide formed a Ca(2+)-dependent and hydroxylamine-sensitive (32)P-labeled phosphoprotein of 110 kDa in the presence of [gamma-(32)P]ATP. Cyclopiazonic acid, but not thapsigargin, blocked this phosphoprotein formation. Transgenic parasites expressing constructs of TcSCA with green fluorescent protein exhibited co-localization of TcSCA with the endoplasmic reticulum proteins BiP and calreticulin. An endoplasmic reticulum location was also found in amastigotes and trypomastigotes using a polyclonal antibody against a COOH-terminal region of the protein. The ability of TcSCA to restore growth of mutant pmr1 pmc1 cnb 1 on medium containing Mn(2+) suggests that TcSCA may also regulate Mn(2+) homeostasis by pumping Mn(2+) into the endoplasmic reticulum of T. cruzi.
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PMID:TcSCA complements yeast mutants defective in Ca2+ pumps and encodes a Ca2+-ATPase that localizes to the endoplasmic reticulum of Trypanosoma cruzi. 1138 80

Mammalian BiP/GRP78 and Escherichia coli DnaK belong to the highly conserved hsp70 family and function as molecular chaperones in the endoplasmic reticulum or the cytosol, respectively. Induction of murine BiP/GRP78 expression in E. coli leads to growth arrest and cell death, independent of the bacterial strain and vector used. Analysis of various BiP constructs and mutants shows that the dominant-lethal phenotype is induced specifically by the expression of the 13.7-kDa C-terminal domain and abolished by a single substitution in that region. Deletion of that region also results in nontoxic gene products that can be overexpressed and purified to homogeneity. The nontoxic mutants are highly expressed in E. coli, representing up to 20% of the soluble fraction. They are catalytically active, depolymerize upon binding ATP or synthetic peptide, and interact with the J-domain of the DnaJ-like accessory protein, MTJ1, with near wild-type affinity. Our data indicate that the cytotoxic effect encountered during overexpression of recombinant proteins can be caused by a single domain and can be alleviated by a specific mutation or deletion in that region without altering the catalytic properties of the enzyme.
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PMID:Isolation, expression, and characterization of fully functional nontoxic BiP/GRP78 mutants. 1138 13

Unassembled Ig heavy chains are retained in the ER via the binding of BiP to the C(H)1 domain, which remains unoxidized. Interestingly, this domain folds rapidly, albeit nonproductively, when heavy chains are released from BiP in vitro with ATP. The in vivo cycling of BiP from heavy chains was monitored using BiP ATPase mutants as kinetic traps. Our data suggest that BiP does not cycle from the C(H)1 domain of free heavy chains. However, heavy and light chain assembly occurs rapidly and requires the ATP-dependent release of BiP. We propose that BiP's ATPase cycle is stalled or nonproductive when it is bound to free heavy chains. The binding of light chains to the complex reactivates the cycle and releases BiP.
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PMID:Unassembled Ig heavy chains do not cycle from BiP in vivo but require light chains to trigger their release. 1148 42

Calnexin (CNX) is a membrane protein of the endoplasmic reticulum that has been defined primarily as a lectin, yet is capable of functioning as a molecular chaperone with non-glycosylated proteins in vitro. Here, we assess the relative contributions of the oligosaccharide- and polypeptide-binding sites of CNX to its in vitro chaperone functions by comparing it with the Hsp70 chaperone of the endoplasmic reticulum, BiP. Both proteins were equally effective in preventing the aggregation of non-glycosylated citrate synthase, indicating that the polypeptide-binding site of CNX is capable of functioning at a level similar to that of Hsp70. However, when confronted with glycoprotein substrates, the lectin site of CNX provided a significant advantage over BiP in suppressing aggregation. CNX also cooperated with BiP and the J domain of Sec63p in the ATP-dependent refolding of glycoprotein and non-glycosylated substrates. The lectin site of CNX was essential for refolding of the glycoprotein. These findings reinforce the function of CNX as a bona fide chaperone and illustrate how its lectin site confers advantages relative to other chaperones when confronted with glycoprotein substrates.
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PMID:Relationship between calnexin and BiP in suppressing aggregation and promoting refolding of protein and glycoprotein substrates. 1151 79

Salicylic acid (SA), an endogenous signaling molecule of plants, possesses anti-inflammatory and anti-neoplastic actions in human. Its derivative, aspirin, is the most commonly used anti-inflammatory and analgesic drug. Aspirin and sodium salicylate (salicylates) have been reported to have multiple pharmacological actions. However, it is unclear whether they bind to a cellular protein. Here, we report for the first time the purification from human fibroblasts of a approximately 78 kDa salicylate binding protein with sequence identity to immunoglobulin heavy chain binding protein (BiP). The Kd values of SA binding to crude extract and to recombinant BiP were 45.2 and 54.6 microM, respectively. BiP is a chaperone protein containing a polypeptide binding site recognizing specific heptapeptide sequence and an ATP binding site. A heptapeptide with the specific sequence displaced SA binding in a concentration-dependent manner whereas a control heptapeptide did not. Salicylates inhibited ATPase activity stimulated by this specific heptapeptide but did not block ATP binding or induce BiP expression. These results indicate that salicylates bind specifically to the polypeptide binding site of BiP in human cells that may interfere with folding and transport of proteins important in inflammation.
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PMID:Aspirin and salicylate bind to immunoglobulin heavy chain binding protein (BiP) and inhibit its ATPase activity in human fibroblasts. 1168 71

The maturation of lipoprotein lipase (LPL) into a catalytically active enzyme was believed to occur only after its transport from the endoplasmic reticulum (ER) to the Golgi apparatus. To test this hypothesis, LPL located in these two subcellular compartments was separated and compared. Heparin affinity chromatography resolved low affinity, inactive LPL displaying ER characteristics from a high affinity, active fraction exhibiting both ER and Golgi forms. The latter forms were further separated by beta-ricin chromatography and were found to have comparable activities per unit of LPL mass. Thus, LPL must reach a functional conformation in the ER. Active LPL, regardless of its cellular location, exhibited the expected dimer conformation. However, inactive LPL, found only in the ER, was highly aggregated. Kinetic analysis indicated a concurrent formation of LPL dimer and aggregate and indicated that the two forms have dissimilar fates. Whereas the dimer remained stable even when confined to the ER, the aggregate was degraded. Degradation rates were not affected by proteasomal or lysosomal inhibitors but were markedly reduced by ATP depletion. Lowering the redox potential in the ER by dithiothreitol caused the dimer to associate with calnexin, BiP, and protein-disulfide isomerase to form large, inactive complexes; dithiothreitol removal induced complex dissociation with restoration of the functional LPL dimer. In contrast, the LPL aggregate was only poorly associated with ER chaperones, appearing to be trapped in an irreversible, inactive conformation destined for ER degradation.
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PMID:Maturation of lipoprotein lipase in the endoplasmic reticulum. Concurrent formation of functional dimers and inactive aggregates. 3142 May 23

In mammalian cells, most membrane proteins are inserted cotranslationally into the ER membrane at sites termed translocons. Although each translocon forms an aqueous pore, the permeability barrier of the membrane is maintained during integration, even when the otherwise tight ribosome-translocon seal is opened to allow the cytoplasmic domain of a nascent protein to enter the cytosol. To identify the mechanism by which membrane integrity is preserved, nascent chain exposure to each side of the membrane was determined at different stages of integration by collisional quenching of a fluorescent probe in the nascent chain. Comparing integration intermediates prepared with intact, empty, or BiP-loaded microsomes revealed that the lumenal end of the translocon pore is closed by BiP in an ATP-dependent process before the opening of the cytoplasmic ribosome-translocon seal during integration. This BiP function is distinct from its previously identified role in closing ribosome-free, empty translocons because of the presence of the ribosome at the translocon and the nascent membrane protein that extends through the translocon pore and into the lumen during integration. Therefore, BiP is a key component in a sophisticated mechanism that selectively closes the lumenal end of some, but not all, translocons occupied by a nascent chain. By using collisional quenchers of different sizes, the large internal diameter of the ribosome-bound aqueous translocon pore was found to contract when BiP was required to seal the pore during integration. Therefore, closure of the pore involves substantial conformational changes in the translocon that are coupled to a complex sequence of structural rearrangements on both sides of the ER membrane involving the ribosome and BiP.
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PMID:A new role for BiP: closing the aqueous translocon pore during protein integration into the ER membrane. 1180 91

HSP70 chaperones mediate protein folding by ATP-dependent interaction with short linear peptide segments that are exposed on unfolded proteins. The mode of action of the Escherichia coli homolog DnaK is representative of all HSP70 chaperones, including the endoplasmic reticulum variant BiP/GRP78. DnaK has been shown to be effective in assisting refolding of a wide variety of prokaryotic and eukaryotic proteins, including the alpha-helical homodimeric secretory cytokine interferon-gamma (IFN-gamma). We screened solid-phase peptide libraries from human and mouse IFN-gamma to identify DnaK-binding sites. Conserved DnaK-binding sites were identified in the N-terminal half of helix B and in the C-terminal half of helix C, both of which are located at the IFN-gamma dimer interface. Soluble peptides derived from helices B and C bound DnaK with high affinity in competition assays. No DnaK-binding sites were found in the loops connecting the alpha-helices. The helix C DnaK-binding site appears to be conserved in most members of the superfamily of interleukin (IL)-10-related cytokines that comprises, apart from IL-10 and IFN-gamma, a series of recently discovered small secretory proteins, including IL-19, IL-20, IL-22/IL-TIF, IL-24/MDA-7 (melanoma differentiation-associated gene), IL-26/AK155, and a number of viral IL-10 homologs. These cytokines belong to a relatively small group of homodimeric proteins with highly interdigitated interfaces that exhibit the strongly hydrophobic character of the interior core of a single-chain folded domain. We propose that binding of DnaK to helix C in the superfamily of IL-10-related cytokines may constitute the hallmark of a novel conserved regulatory mechanism in which HSP70-like chaperones assist in the formation of a hydrophobic dimeric "folding" interface.
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PMID:The conserved helix C region in the superfamily of interferon-gamma /interleukin-10-related cytokines corresponds to a high-affinity binding site for the HSP70 chaperone DnaK. 1197 Sep 58

BiP is an Hsp70 homologue found in the endoplasmic reticulum of eukaryotic cells. Like other Hsp70 chaperones, BiP interacts with its substrate proteins in an ATP-dependent manner. The functional analysis has so far been performed mainly with short, synthetic peptides. Here, we present an experimental system that allows to study the partial reactions of the BiP chaperone cycle for a natural substrate protein domain in its soluble, stably unfolded conformation. This unfolded antibody domain forms a binary complex with BiP in the absence of ATP. The dissociation of the BiP dimer seems to be the rate-limiting step in this reaction. The BiP-C(H)3 complexes dissociate rapidly in the presence of ATP. The affinity for BiP-binding peptides and the non-native antibody domain was determined to be similar, suggesting that only the peptide binding site is involved in these interactions. Furthermore, these results imply that, also in the context of the antibody domain, an extended peptide sequence is recognized. However, the accessibility of the BiP-binding site in the non-native protein seems to influence the kinetics of complex formation.
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PMID:Interaction of the chaperone BiP with an antibody domain: implications for the chaperone cycle. 1205 9


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