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)

Glycosylation plays a crucial role in glycoprotein stability and its correct folding. Murine acid sphingomyelinase (ASM) is a lysosomal glycoprotein. We studied the functional role of its individual N-linked oligosaccharides needed to maintain enzymatic activity and protein stability. Mutagenized cDNA constructs were heterologously expressed. All six potential N-glycosylation sites were modified. Incomplete glycosylation of the most distant C-terminal site resulted in two isoforms. Oligosaccharides at N-84, N-173, and N-611 were found to be of minor importance for enzymatic activity. The glycosylation defect at N-333 or N-393 reduced the enzymatic activity to 40% and at N-518 to less than 20%. These mutations did not effect the Km value. Glycosylation at N-333 and N-393 mainly contributed to the enzyme stability and prevented degradation at lysosomal acidic pH, whereas the low residual enzymatic activity of mutant ASM deficient in glycosylation at N-518 was caused by protein misfolding. The mutant protein was also prone to proteolysis when trapped in the endoplasmic reticulum/cis-Golgi after brefeldin A application. Insufficiently glycosylated ASM formed a stable complex with BiP, an immunoglobulin heavy chain-binding protein, and thus remained in the endoplasmic reticulum. 32PO4 labeling revealed that the glycosylation mutants of ASM were phosphorylated predominantly at mannose residues of oligosaccharides linked to N-84, N-333, and N-393.
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PMID:Functional analysis of the glycosylation of murine acid sphingomyelinase. 894 61

Herein we describe detailed characterization of four common mutations (L302P, H421Y, R496L and DeltaR608) within the acid sphingomyelinase (ASM) gene causing types A and B Niemann-Pick disease (NPD). In vitro and in situ enzyme assays revealed marked deficiencies of ASM activity in NPD cell lines homoallelic for each mutation, although Western blotting and fluorescent microscopy showed that the mutant ASM polypeptides were expressed at normal levels and trafficked to lysosomes. Co-immunoprecipitation of the polypeptides with the ER chaperone, BiP, confirmed these findings, as did in vitro expression of the mutant cDNAs in reticulocyte lysates. We further developed a computer assisted, three-dimensional model of human ASM based on homologies to known proteins, and used this model to map each NPD mutation in relation to putative substrate binding, hydrolysis and zinc-binding domains. Lastly, we generated transgenic mice expressing the R496L and DeltaR608 mutations on the complete ASM knock-out background (ASMKO), and established breeding colonies for the future evaluation of enzyme enhancement therapies. Analysis of these mice demonstrated that the mutant ASM transgenes were expressed at high levels in the brain, and in the case of the DeltaR608 mutation, produced residual ASM activity that was significantly above the ASMKO background.
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PMID:Characterization of common SMPD1 mutations causing types A and B Niemann-Pick disease and generation of mutation-specific mouse models. 1881 62

Melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) uniquely displays broad cancer-specific apoptosis-inducing activity through induction of endoplasmic reticulum (ER) stress. We hypothesize that ceramide, a promoter of apoptosis, might contribute to mda-7/IL-24 induction of apoptosis. Ad.mda-7-infected tumor cells, but not normal cells, showed increased ceramide accumulation. Infection with Ad.mda-7 induced a marked increase in various ceramides (C16, C24, C24:1) selectively in prostate cancer cells. Inhibiting the enzyme serine palmitoyltransferase (SPT) using the potent SPT inhibitor myriocin (ISP1), impaired mda-7/IL-24-induced apoptosis and ceramide production, suggesting that ceramide formation caused by Ad.mda-7 occurs through de novo synthesis of ceramide and that ceramide is required for mda-7/IL-24-induced cell death. Fumonisin B1 (FB1) elevated ceramide formation as well as apoptosis induced by Ad.mda-7, suggesting that ceramide formation may also occur through the salvage pathway. Additionally, Ad.mda-7 infection enhanced expression of acid sphingomyelinase (ASMase) with a concomitant increase in ASMase activity and decreased sphingomyelin in cancer cells. ASMase silencing by RNA interference inhibited the decreased cell viability and ceramide formation after Ad.mda-7 infection. Ad.mda-7 activated protein phosphatase 2A (PP2A) and promoted dephosphorylation of the anti-apoptotic molecule BCL-2, a downstream ceramide-mediated pathway of mda-7/IL-24 action. Pretreatment of cells with FB1 or ISP-1 abolished the induction of ER stress markers (BiP/GRP78, GADD153 and pospho-eIF2alpha) triggered by Ad.mda-7 infection indicating that ceramide mediates ER stress induction by Ad.mda-7. Additionally, recombinant MDA-7/IL-24 protein induced cancer-specific production of ceramide. These studies define ceramide as a key mediator of an ER stress pathway that may underlie mda-7/IL-24 induction of cancer-specific killing.
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PMID:Ceramide plays a prominent role in MDA-7/IL-24-induced cancer-specific apoptosis. 1993 35