Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UMLS:C0155339 (Brown)
12,436 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have raised two monospecific antibodies against synthetic peptides derived from the membrane domain of the ER glycoprotein 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the rate limiting enzyme in the cholesterol biosynthetic pathway. This domain, which was proposed to span the ER membrane seven times (Liscum, L., J. Finer-Moore, R. M. Stroud, K. L. Luskey, M. S. Brown, and J. L. Goldstein. 1985. J. Biol. Chem. 260:522-538), plays a critical role in the regulated degradation of the enzyme in the ER in response to sterols. The antibodies stain the ER of cells and immunoprecipitate HMG-CoA reductase and HMGal, a chimeric protein composed of the membrane domain of the reductase fused to Escherichia coli beta-galactosidase, the degradation of which is also accelerated by sterols. We show that the sequence Arg224 through Leu242 of HMG-CoA reductase (peptide G) faces the cytoplasm both in cultured cells and in rat liver, whereas the sequence Thr284 through Glu302 (peptide H) faces the lumen of the ER. This indicates that a sequence between peptide G and peptide H spans the membrane of the ER. Moreover, by epitope tagging with peptide H, we show that the loop segment connecting membrane spans 3 and 4 is sequestered in the lumen of the ER. These results demonstrate that the membrane domain of HMG-CoA reductase spans the ER eight times and are inconsistent with the seven membrane spans topological model. The approximate boundaries of the proposed additional transmembrane segment are between Lys248 and Asp276. Replacement of this 7th span in HMGal with the first transmembrane helix of bacteriorhodopsin abolishes the sterol-enhanced degradation of the protein, indicating its role in the regulated turnover of HMG-CoA reductase within the endoplasmic reticulum.
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PMID:Immunological evidence for eight spans in the membrane domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase: implications for enzyme degradation in the endoplasmic reticulum. 137 17

Mutant V.24.1, a member of the End4 complementation group of temperature-sensitive CHO cells, is defective in secretion at the restrictive temperature (Wang, R.-H., P. A. Colbaugh, C.-Y. Kao, E. A. Rutledge, and R. K. Draper. 1990. J. Biol. Chem. 265:20179-20187; Presley, J. F., R. K. Draper, and D. T. Brown. 1991. J. Virol. 65:1332-1339). We have further investigated the secretory lesion and report three main findings. First, the block in secretion is not due to aberrant folding or oligomerization of secretory proteins in the endoplasmic reticulum because the hemagglutinin of influenza virus folded and oligomerized at the same rate in mutant and parental cells at the restrictive temperature. Second, secretory proteins accumulated in a compartment intermediate between the ER and the Golgi. Several lines of evidence support this conclusion, the most direct being the colocalization by immunofluorescence microscopy of influenza virus hemagglutinin with a 58-kD protein that is known to reside in an intermediate compartment. Third, at the resolution of fluorescence microscopy, the Golgi complex in the mutant cells vanished at the restrictive temperature.
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PMID:Retention of secretory proteins in an intermediate compartment and disappearance of the Golgi complex in an END4 mutant of Chinese hamster ovary cells. 157 51

The influenza virus hemagglutinin (HA) temperature-sensitive (ts) mutant, ts61S, contains a nucleotide change in RNA segment 4 which leads to an amino acid change at HA1 residue 110 of serine to proline. When ts61S HA is synthesized and maintained at the nonpermissive temperature (39.5 degrees), the HA is defective in transport in the exocytic pathway and is retained in the endoplasmic reticulum (S. Nakajima, D. J. Brown, M., Ueda, K., Nakajima, A. Suguira, A. K. Pattnaik, and D. P. Nayak, 1986, Virology 154, 279-285). In a comparison of the biochemical properties of ts61S HA and A/WSN/33 HA (wt) expressed at the permissive temperature (33 degrees), we have found that ts61S HA is extensively debilitated. A large proportion of ts61S HA fails to gain reactivity with conformation-specific monoclonal antibodies and does not become resistant to protease digestion. In turn, a large population of the molecules are not transported from the ER to the Golgi apparatus or cell surface with the same kinetics or efficiency as wt HA. These data suggest that the serine to proline change at HA1 residue 110 leads to partial impairment of folding at the permissive temperature with complete impairment at the nonpermissive temperature.
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PMID:Influenza virus ts61S hemagglutinin is significantly defective in polypeptide folding and intracellular transport at the permissive temperature. 192 89

The differentiation of brown adipocyte precursor cells was studied in interscapular brown adipose tissue of adult mice by electron microscopy. Different stages of cell differentiation were characterized in situ. Previous autoradiographic studies suggested that interstitial cells represent the precursor cells of fully differentiated brown adipocytes. The present observations provide morphological evidence for a progressive differentiation of interstitial stem cells into mature brown adipocytes. Four typical stages of development were identified: (1) interstitial cells, (2) protoadipocytes, (3) preadipocytes, and (4) mature brown adipocytes. Interstitial stem cells were small spindle shaped cells, situated between brown adipocytes and characterized by a high nuclear-cytoplasmic ratio, the scarcity of organelles, and the absence of lipid inclusions. Protoadipocytes resembled interstitial cells except that they contained a few tiny lipid droplets in their cytoplasm. Preadipocytes had a larger cytoplasm enclosing many mitochondria and lipid droplets; the smooth endoplasmic reticulum was well developed surrounding the lipid droplets, and was closely associated with the mitochondria. Preadipocytes had the typical structure of growing cells, developing long cytoplasmic processes between and around blood capillaries. Mature brown adipocytes represented the final stage of differentiation. Almost all their cellular volume was occupied by lipid droplets and numerous mitochondria with very dense cristae. Brown adipocytes were also characterized by a tight association with blood capillaries, as expected from metabolically active cells requiring oxygen and substrates. These observations provide direct ultrastructural evidence for a progressive differentiation of interstitial cells into brown adipocytes with a continuum of intermediate cellular types.
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PMID:In vivo differentiation of brown adipocytes in adult mice: an electron microscopic study. 239 93

The ligand-binding domain of the low density lipoprotein (LDL) receptor is composed of seven cysteine-rich repeats, each approximately 40 amino acids long. Previous studies by van Driel et al. [van Driel, I. R., Goldstein, J. L., Sudhof, T. C. & Brown, M. S. (1987) J. Biol. Chem. 262, 17443-17449] showed that if the first repeat of the ligand-binding domain (encoded by exon 2) is deleted, the receptor fails to bind an anti-LDL receptor monoclonal antibody (IgG-C7) but continues to bind LDL with high affinity. Cultured fibroblasts from a Black South African Xhosa patient (TT) with the clinical syndrome of homozygous familial hypercholesterolemia demonstrated high-affinity cell-surface binding of 125I-labeled LDL but not 125I-labeled IgG-C7. Previous haplotype analysis, using 10 restriction fragment length polymorphic sites, suggested that the patient inherited two identical LDL receptor alleles. The polymerase chain reaction technique was used to selectively amplify exon 2 of the LDL receptor gene from this patient. Sequence analysis of the amplified fragment disclosed a deletion of six base pairs that removes two amino acids, aspartic acid and glycine, from the first cysteine-rich ligand binding repeat. The mutation creates a new PstI restriction site that can be used to detect the deletion. The existence of this mutant allele confirms that the epitope of IgG-C7 is located in the first cysteine-rich repeat and that this repeat is not necessary for LDL binding. The mutant gene produced a normally sized 120-kilodalton LDL receptor precursor protein that matured to the 160-kilodalton form at less than one-fourth the normal rate. Thus, deletion of two amino acids within the first cysteine-rich repeat retards receptor transport from the endoplasmic reticulum to the cell surface, in contrast to deletion of the entire first repeat, which has no effect on receptor maturation.
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PMID:Deletion in the first cysteine-rich repeat of low density lipoprotein receptor impairs its transport but not lipoprotein binding in fibroblasts from a subject with familial hypercholesterolemia. 326 45

Brown adipose tissue uncoupling protein (UCP), an integral polytopic protein of the mitochondrial inner membrane, is composed of at least six transmembrane segments whose net hydrophobic character derives from paired amphiphilic helices. The protein is synthesized in the cytoplasm as a polypeptide (307 amino acids) lacking a cleavable targeting (signal) peptide. Deletion mutagenesis and fusion protein constructions revealed the existence of at least two import signals: one lying between UCP precursor amino acids 13-105 and the other downstream of position 101. The former resulted in both targeting and membrane insertion of a fusion protein, whereas the latter targeted UCP 102-307 into the organelle but failed to result in membrane insertion. When a strong matrix-targeting signal derived from precarbamoyl phosphate synthetase was fused to UCP amino acids 169-307 or 52-307 (containing three and five transmembrane domains, respectively), the fusion proteins were efficiently imported to the soluble matrix compartment where correct signal cleavage took place. We suggest that assembly of UCP into the inner membrane follows a coordinate insertion pathway for integration and may use more than one signal sequence to achieve this. In this respect, it might share certain mechanistic features with the insertion of polytopic proteins into the endoplasmic reticulum. The data also suggest, however, that integration of the amino-terminal third of UCP into the inner membrane may be required to help or enhance insertion of the remaining UCP transmembrane domains.
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PMID:Topogenesis of mitochondrial inner membrane uncoupling protein. Rerouting transmembrane segments to the soluble matrix compartment. 341 58

Adipocytes in intact brown adipose tissue show multivacuolar lipid deposit and mitochondria of 'typical' morphology. Cultured brown adipocytes retain the multivacuolar lipid deposit, while 'typical' mitochondria degenerate and 'atypical' organelles appear instead of the former. Since evidence exists that catecholamines deeply influence brown adipose tissue morphology and function in vivo, we undertook the present ultrastructural investigation to assess whether exposure of cultured brown fat cell to noradrenaline could prevent (or induce regression of) the in vitro morphological modifications of brown adipocytes. Brown adipocytes cultured for 8 h in the presence of noradrenaline (5 X 10(-5) M) had a larger mitochondrial area (i.e. a larger percentage of cytoplasm occupied by non-degenerating mitochondria) in comparison with control cells, as assessed by morphometry; this was due to larger number of mitochondria in noradrenaline-treated cells. Moreover, a number of cells with mitochondria strictly resembling those of the intact tissue were visible in noradrenaline-treated cultured after 8 hr, while 'typical' mitochondria were no longer observed in parallel control cultures. After 5 days of culture without hormone addition, exposure to noradrenaline (5 X 10(-5) M) did not induce quantitative modifications of 'atypical' mitochondria or changes of their ultrastructure up to 12 hr. However, reduction in size of the lipid deposit and activation of both rough endoplasmic reticulum and Golgi apparatus were evident in noradrenaline-treated adipocytes in comparison with non-treated cells.
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PMID:Effects of noradrenaline exposure on rat brown adipocytes in cultures. An ultrastructural study. 343 11

The clearance of many oxidized drugs falls with age. Whilst factors such as reduced liver size, blood flow and specific enzyme activity may be important, the possibility that reduced enzyme affinity for substrate contributes to this fall has not hitherto been investigated. Using liver microsomes from 12 young adult and 12 elderly male Norwegian Brown rats we defined the kinetics of ethoxyresorufin-O-de-ethylation and aldrin epoxidation, specific substrates for the 3-methylcholanthrene inducible and phenobarbitone inducible forms of cytochrome P450, respectively. Our results show a marked fall in the maximal activity of both enzymes in advanced age whether expressed in terms of microsomal protein or unit of cytochrome P450, but with no change in apparent enzyme affinity (Km). Since Km is unchanged, we feel that qualitative age-related changes in cytochrome P450 are unlikely. Reduced metabolism may be due to age-related alterations in coenzymes or smooth endoplasmic reticulum lipid membranes.
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PMID:The effect of age on mono-oxygenase enzyme kinetics in rat liver microsomes. 360 96

The microsomal enzyme 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase is subject to rapid degradation when cells are incubated with sterols or mevalonic acid (MVA). It has been shown that this rapid degradation is dependent upon both a sterol and another MVA-derived metabolite (Nakanishi, M., Goldstein, J. L., and Brown, M. S. (1988) J. Biol. Chem. 258, 8929-8937). In the current study, inhibitors of the isoprene biosynthetic pathway were used to define further this mevalonic acid derivative involved in the accelerated degradation of HMG-CoA reductase. The accelerated degradation of HMG-CoA reductase in met-18b-2 cells, which is induced by the addition of MVA, was inhibited by the presence of the squalene synthase inhibitor, zaragozic acid/squalestatin, or the squalene epoxidase inhibitor, NB-598. Accelerated degradation of HMG-CoA reductase was observed when NB-598-treated cells were incubated with both MVA and sterols. In contrast, the addition of MVA and sterols to zaragozic acid/squalestatin-treated cells did not result in rapid enzyme degradation. This MVA- and sterol-dependent degradation of HMG-CoA reductase persisted in cells permeabilized with reduced streptolysin O. Finally, the selective degradation of HMG-CoA reductase was also observed in rat hepatic microsomes incubated in vitro in the absence of ATP and cytosol. We conclude that the MVA-derived component that is required for the accelerated degradation of HMG-CoA reductase is derived from farnesyl disphosphate and/or squalene in the isoprenoid biosynthetic pathway. We propose that this component has a permissive effect and does not, by itself, induce the degradation of HMG-CoA reductase. We also conclude that the degradation of HMG-CoA occurs in the endoplasmic reticulum, and, once the degradation of HMG-CoA reductase has been initiated by MVA and sterols, all necessary components for the continued degradation of HMG-CoA reductase reside in the endoplasmic reticulum.
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PMID:Mevalonic acid-dependent degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in vivo and in vitro. 827 63

The rigidly ordered icosahedral lattice of the Sindbis virus envelope is composed of a host-derived membrane bilayer in which the viral glycoproteins E1 and E2 reside. E1-E1 interactions stabilized by intramolecular disulfide bridges play a significant role in maintaining the envelope's structural integrity (R. P. Anthony and D. T. Brown, J. Virol. 65:1187-1194, 1991; R. P. Anthony, A. M. Paredes, and D. T. Brown, Virology 190:330-336, 1992). We have examined the acquisition of disulfide bridges within E1 during its maturation. Prior to exit from the endoplasmic reticulum, E1 folds via at least three intermediates, differing in the number and/or arrangement of their disulfides, into a single, compact form. This E1 species remains stable with respect to its disulfides until late in the secretory pathway, when E1 attains a metastable conformation. At this point, when appropriately triggered, intramolecular thiol-disulfide exchange reactions within E1 can occur, resulting in the generation of alternative E1 species. This metastable nature of mature E1 may have important implications for the mechanism of virus disassembly during the initial stages of the infection process (B. Abell and D. T. Brown, J. Virol. 67:5496-5501, 1993).
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PMID:Formation and rearrangement of disulfide bonds during maturation of the Sindbis virus E1 glycoprotein. 828 84


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