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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The substructural organization of completely crystalline peroxisomes present in Hansenula polymorpha cells grown under methanol limitation in a chemostat was investigated by different cytochemical and ultrastructural techniques. Time-dependent cytochemical staining experiments indicated that activities of the two main constituents of these organelles, namely, alcohol oxidase and catalase, were present throughout the crystalline matrix. Catalase was completely removed from isolated peroxisomes by osmotic shock treatment. After such treatment, the ultrastructure of the crystalline matrix of the organelles remained virtually intact. Because alcohol oxidase activity was still present in this matrix, it was concluded that alcohol oxidase protein is the only structural element of the peroxisomal crystalloids. The molecular architecture of the crystalloids was investigated in ultrathin cryosections which permitted recognition of individual molecules in the crystalline matrix. Depending on the plane of sectioning, different crystalline patterns were observed. Tilting experiments indicated that these images were caused by superposition of octameric alcohol oxidase molecules arranged in a tetragonal lattice. A three-dimensional model of the crystalloid is presented. The repeating unit of this structure is composed of four alcohol oxidase molecules. The crystalloid represents an open structure, which may explain the observed free mobility of catalase molecules.
Mol Cell Biol 1981 Oct
PMID:Substructure of crystalline peroxisomes in methanol-grown Hansenula polymorpha: evidence for an in vivo crystal of alcohol oxidase. 705 Jun 59

The kinetics of secretion of proteins by Leishmania braziliensis was followed by incorporation of [3H]leucine into macromolecules produced by the cells which are released into the growth medium. About 10% of the total protein synthesized by actively growing cells is secreted. Cycloheximide (100 microgram/ml) and puromycin (0.5 mM) inhibited the incorporation of labelled leucine by 85 and 99%, respectively. The secreted proteins do not seem to result from cell lysis since, first, the kinetics of production are linear and, secondly, less than 1% of thymidine or uridine incorporated by the cells is found in the medium. Cells grown with [3H]leucine and then transferred to fresh medium show two phases of secretion. During the first six hours, it is slow and reaches a plateau. The release increases about ten-fold during the next six hours. An analysis of the secreted material showed that following precipitation with methanol and sodium acetate, three isotopically labelled peaks were eluted from Sephadex G-120-150. The first of these, containing 50% of the radioactivity, did not react with anti-leishmanial serum, while the last two did. Since the last two fractions could be labelled with [3H]glucosamine as well as [3H]leucine it is suggested that they are glycoprotein in nature and are similar to the products released by other species of Leishmania.
Mol Biochem Parasitol 1980 Jun
PMID:Production and secretion of Leishmania braziliensis proteins. 744 14

To understand the role of N-acetylaspartate (NAA) as an acetyl donor, we investigated the metabolism of NAA in brain and liver slice preparations. The tissue slices were incubated with [14C-acetyl]NAA (SA = 3 microCi/mumol) or [14C]acetate (SA = 3 microCi/mumol) for 2 h. The tissue was homogenized and was extracted using chloroform/methanol (2:1). The aqueous phase was initially analyzed using anion exchange HPLC while the lipid phase was analyzed using a two-dimensional TLC system. Further resolution of the NAA peak from the anion exchange HPLC was performed using a reverse phase HPLC system. The aqueous phase of both the liver and brain samples incubated with [14C-acetyl]NAA revealed similar patterns of three distinct radioactivity peaks corresponding to NAA, acetate and an early eluting unknown molecule. Further resolution of the NAA peak using reverse phase HPLC indicated that it corresponded to NAA and acetyl CoA. There was significant incorporation of radioactivity into various lipid components in both the brain and liver samples. Patterns similar to that observed with NAA were detected in the case of [14C]acetate in both the brain and liver slice preparations. These results demonstrate that NAA metabolism is not restricted to the nervous system, although its biosynthesis is. It is clear that acetyl moiety of NAA is incorporated into lipids and partially hydrolyzed to free acetate in both brain and liver preparations. Further, production of acetyl CoA from NAA indicates that the acetyl group of NAA is incorporated into lipids and perhaps other acetylated molecules via the acetyl CoA route. A working hypothesis on the metabolic role of NAA is presented.
Brain Res Mol Brain Res 1995 Jul
PMID:N-acetylaspartate as an acetyl source in the nervous system. 747 23

African swine fever virus polypeptides with molecular weight of 120, 78, 69, 59, 56, 45, 39, 28, 26, 24, 16, and 14 kD are the major proteins in the purified virions, as shown by electrophoresis and immunoblotting. A mixture of proteases and pancreatic lipase hydrolyzed the polypeptides of 120 and 78 kD in viral preparations at low concentrations of enzymes, polypeptides of 69, 56, 45, 39, 28, and 14 kD disappeared after treatment with this mixture at medium concentrations, and 26 kD polypeptide was eliminated at a high concentration of the enzymes. The 21 kD polypeptide which did not react with the specific antiviral serum in immunoblotting was not hydrolyzed by proteases contaminating lipase. Treatment with triton X-100 and ether boosted the activity of DNA-dependent RNA-polymerase, whereas treatment with ether followed by resedimentation markedly decreased polymerase activity in the resultant sediment. Treatment with diethyl ether did not influence the activity of virus-associated ATPase, which was partially resistant to denaturating organic solvents acetone and chloroform-methanol mixture. Our findings and published data permitted us to propose a schematic arrangement of viral polypeptides and enzymes in the virion structure.
Mol Gen Mikrobiol Virusol
PMID:[Localizing the major peptides of African swine fever virus and virus-associated enzymes in the virion structure]. 747 38

The serine/threonine kinase Raf-1 functions downstream from Ras to activate mitogen-activated protein kinase kinase, but the mechanisms of Raf-1 activation are incompletely understood. To dissect these mechanisms, wild-type and mutant Raf-1 proteins were studied in an in vitro system with purified plasma membranes from v-Ras- and v-Src-transformed cells (transformed membranes). Wild-type (His)6- and FLAG-Raf-1 were activated in a Ras- and ATP-dependent manner by transformed membranes; however, Raf-1 proteins that are kinase defective (K375M), that lack an in vivo site(s) of regulatory tyrosine (YY340/341FF) or constitutive serine (S621A) phosphorylation, that do not bind Ras (R89L), or that lack an intact zinc finger (CC165/168SS) were not. Raf-1 proteins lacking putative regulatory sites for an unidentified kinase (S259A) or protein kinase C (S499A) were activated but with apparently reduced efficiency. The kinase(s) responsible for activation by Ras or Src may reside in the plasma membrane, since GTP loading of plasma membranes from quiescent NIH 3T3 cells (parental membranes) induced de novo capacity to activate Raf-1. Wild-type Raf-1, possessing only basal activity, was not activated by parental membranes in the absence of GTP loading. In contrast, Raf-1 Y340D, possessing significant activity, was, surprisingly, stimulated by parental membranes in a Ras-independent manner. The results suggest that activation of Raf-1 by phosphorylation may be permissive for further modulation by another membrane factor, such as a lipid. A factor(s) extracted with methanol-chloroform from transformed membranes or membranes from Sf9 cells coexpressing Ras and SrcY527F significantly enhanced the activity of Raf-1 Y340D or active Raf-1 but not that of inactive Raf-1. Our findings suggest a model for activation of Raf-1, wherein (i) Raf-1 associates with Ras-GTP, (ii) Raf-1 is activated by tyrosine and/or serine phosphorylation, and (iii) Raf-1 activity is further increased by a membrane cofactor.
Mol Cell Biol 1995 Aug
PMID:Regulation of Raf-1 and Raf-1 mutants by Ras-dependent and Ras-independent mechanisms in vitro. 762 7

Extensive experimental data are available on the native, partially and fully unfolded states of ubiquitin. Two and three-dimensional NMR experiments of a partially unfolded form of the protein in 60% methanol indicate that approximately one-half of the molecule contains disrupted but native-like structure while the other half is unstructured and/or contains non-native structure. In contrast, the interpretation of hydrogen-exchange data have led to the conclusion that this state is native-like. Thus, there are discrepancies between the experimental studies, or interpretations based on the data. We compare the results of molecular dynamics simulations, under varying conditions, with the experimental results. The simulations extend past 0.5 ns and include explicit solvent molecules: either pure water or 60% methanol. To begin with, ubiquitin was thermally denatured in water (at 498 K). Two particular structures, or "aliquots", during the unfolding process were selected for further study (60 and 198 ps). These structures were then simulated separately in water and 60% methanol at a lower and experimentally meaningful temperature (335 K). The conformations generated from the structure extracted later in the simulation contained significant amounts of non-native structure in the presence of methanol while satisfying both the NMR and hydrogen exchange data. In fact, clearly non-native regions of the structure yielded the desired protection from hydrogen exchange. In contrast, an earlier, more native-like, intermediate did not do as well at predicting the hydrogen-exchange behavior and was inconsistent with the NMR data. These data suggest that the results and interpretations using the different experimental techniques can be reconciled by a single state. This finding also brings into question the practice of interpreting protection to hydrogen exchange in terms of native secondary and tertiary structure, especially when one has weak patterns and low protection factors. When the partially unfolded states were placed in pure water, the protein collapsed and began to refold. Therefore, the desired solvent-dependent properties were observed: the partially unfolded conformations with increased exposure of hydrophobic residues remained expanded in methanol but collapsed in water as the non-polar groups minimized their exposure to solvent.
J Mol Biol 1995 Mar 31
PMID:Molecular dynamics simulations of protein unfolding and limited refolding: characterization of partially unfolded states of ubiquitin in 60% methanol and in water. 771 3

A series of three molecular dynamics simulations at 300 K in explicit solvent environments of chloroform, methanol and water has been performed on the pulmonary surfactant lipoprotein, SP-C, comprising several consecutive valine residues in order to investigate the stability of the alpha-helical conformation. Two additional simulations were performed on truncated SP-C with a five-residue N-terminal deletion at 300 K and 500 K in water, the high temperature run in order to increase the rate of peptide denaturation. Indications of destabilization appear in chloroform during 1 ns while the SP-C alpha-helix is remarkably stable during 1 ns in methanol and water. In particular the polyvalyl part comprising residues Val15 to Val21 remains intact even at elevated temperature, and the valines do not disrupt the alpha-helical conformation. The valyl-rotamer sampling is partly restricted. Unfolding takes place successively along the primary sequence starting from the C-terminal end. Factors affecting polypeptide stability in molecular dynamics simulations are addressed. The intrinsic helix-forming tendency of valine residues and its dependence on the sequence context, and the role of the solvent environment in stabilizing or destabilizing an alpha-helical fold, are discussed.
J Mol Biol 1995 Apr 07
PMID:The effect of environment on the stability of an integral membrane helix: molecular dynamics simulations of surfactant protein C in chloroform, methanol and water. 772 32

Adriamycin (AD)-Fe3+ caused the inactivation of Na(+)-, K(+)-ATPase and Ca(2+)-ATPase of erythrocyte membranes during lipid peroxidation. AD-Fe3+ also induced the formation of fluorescent substances from the membranes with lipid peroxidation. The fluorescent substances were little extracted by chloroform-methanol, indicating that they were retained in the membranes. Butylated hydroxytoluene and trolox strongly inhibited both the inactivation of these ATPases and the formation of fluorescent substances with lipid peroxidation. Another antioxidant, vitamin E, slightly prevented the damage of the membranes. However, p-nitrophenyl phosphatase activity and acetylcholine esterase have lower or no susceptibility to the membrane lipid peroxidation. These results indicated that the ATPases were very sensitive to lipid peroxidation and that the membranes were modified during the peroxidation reaction.
Res Commun Mol Pathol Pharmacol 1995 Feb
PMID:Adriamycin-Fe(3+)-induced inactivation of enzymes in erythrocyte membranes during lipid peroxidation. 774 51

The ability of breast tumours to synthesize hormones is well recognized, and local production of sex steroids is thought to play a role in breast cancer growth. We measured the intratumour and circulating levels of testosterone, dihydrotestosterone (DHT) and oestradiol in 35 histologically confirmed carcinomatous mammary tissues obtained at breast surgery from 34 postmenopausal patients, age 50-85 years. Intra-tissue steroids were extracted with ethanol:acetone (1:1; v/v), defatted with 70% methanol in water, and extracted with ether. Steroids, from tissue and serum, were separated by partition chromatography on celite columns and were measured by RIA. Intratumour testosterone and DHT concentrations were significantly correlated, after the exclusion of an outlier (rs = 0.71; P = 0.0001). No association was found between oestradiol and either of the two androgens. Mean oestradiol and DHT concentrations were significantly higher in tissue than in blood (P = 0.0001). Mean testosterone levels in tissues did not significantly differ from those measured in blood. Our data suggest that at least a part of intratissue DHT is produced locally from testosterone. The meaning of high oestradiol and DHT levels in cancer tissue still needs to be defined.
J Steroid Biochem Mol Biol 1995 Jun
PMID:Testosterone, dihydrotestosterone and oestradiol levels in postmenopausal breast cancer tissues. 777 58

In water-methanol and water-dimethylformamide (DMF) (1:1 v/v) solutions tryptophanase from E.coli retains its abilities to form a quinonoid complex with quasisubstrates and to catalyze the decomposition of S-o-nitrophenyl-L-cysteine (SOPC). Both the KM and Vmax values decrease in water-organic media. The affinities of tryptophanase for L-alanine, L-tryptophan, oxindolyl-L-alanine and indole in aqueous methanol are decreased, the effect being stronger for the more hydrophobic substances. In a water solution tryptophanase catalizes the reaction of SOPC with indole to form L-tryptophan while in water-organic solvents only decomposition of SOPC is observed.
Biochem Mol Biol Int 1994 Aug
PMID:Tryptophanase from Escherichia coli: catalytic and spectral properties in water-organic solvents. 784 21


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