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Protein Processing (protein + processing)
Selected AbstractsA1 Adenosine Receptors Accumulate in Neurodegenerative Structures in Alzheimer's Disease and Mediate Both Amyloid Precursor Protein Processing and Tau Phosphorylation and TranslocationBRAIN PATHOLOGY, Issue 4 2003Ester Angulo Immunostaining of adenosine receptors in the hippocampus and cerebral cortex from necropsies of Alzheimer's disease (AD) patients shows that there is a change in the pattern of expression and a redistribution of receptors in these brain areas when compared with samples from controls. Adenosine A1 receptor (A1R) immunoreactivity was found in degenerating neurons with neurofibrillary tangles and in dystrophic neurites of senile plaques. A high degree of colocalization for A1R and pA4 amyloid in senile plaques and for A1R and tau in neurons with tau deposition, but without tangles, was seen. Additionally, adenosine A2A receptors, located mainly in striatal neurons in controls, appeared in glial cells in the hippocampus and cerebral cortex of patients. On comparing similar samples from controls and patients, no significant change was evident for metabotropic glutamate receptors. In the human neuroblastoma SH-SY5Y cell line, agonists for A1R led to a dose-dependent increase in the production of soluble forms of amyloid precursor protein in a process mediated by PKC. A1R agonist induced p21 Ras activation and ERK1/2 phosphorylation. Furthermore, activation of A1R led to and ERK-dependent increase of tau phosphorylation and translocation towards the cytoskeleton. These results indicate that adenosine receptors are potential targets for AD. [source] Key Factors in Alzheimer's Disease: ,-amyloid Precursor Protein Processing, Metabolism and Intraneuronal TransportBRAIN PATHOLOGY, Issue 1 2001Thomas A. Bayer During the last years it has become evident that the ,-amyloid (A,) component of senile plaques may be the key molecule in the pathology of Alzheimer's disease (AD). The source and place of the neurotoxic action of A,, however, is still a matter of controversy. The precursor of the ,-amyloid peptide is the predominantly neuronal ,-amyloid precursor protein. We, and others, hypothesize that intraneuronal misregulation of APP leads to an accumulation of A, peptides in intracellular compartments. This accumulation impairs APP trafficking, which starts a cascade of pathological changes and causes the pyramidal neurons to degenerate. Enhanced A, secretion as a function of stressed neurons and remnants of degenerated neurons provide seeds for extracellular A, aggregates, which induce secondary degenerative events involving neighboring cells such as neurons, astroglia and macrophages/microglia. [source] Gene expression profiles of lens regeneration and development in Xenopus laevisDEVELOPMENTAL DYNAMICS, Issue 9 2009Erica L. Malloch Abstract Seven hundred and thirty-four unique genes were recovered from a cDNA library enriched for genes up-regulated during the process of lens regeneration in the frog Xenopus laevis. The sequences represent transcription factors, proteins involved in RNA synthesis/processing, components of prominent cell signaling pathways, genes involved in protein processing, transport, and degradation (e.g., the ubiquitin/proteasome pathway), matrix metalloproteases (MMPs), as well as many other proteins. The findings implicate specific signal transduction pathways in the process of lens regeneration, including the FGF, TGF-beta, MAPK, Retinoic acid, Wnt, and hedgehog signaling pathways, which are known to play important roles in eye/lens development and regeneration in various systems. In situ hybridization revealed that the majority of genes recovered are expressed during embryogenesis, including in eye tissues. Several novel genes specifically expressed in lenses were identified. The suite of genes was compared to those up-regulated in other regenerating tissues/organisms, and a small degree of overlap was detected. Developmental Dynamics 238:2340,2356, 2009. © 2009 Wiley-Liss, Inc. [source] A hypermorphic mouse Gli3 allele results in a polydactylous limb phenotypeDEVELOPMENTAL DYNAMICS, Issue 3 2007Chengbing Wang Abstract Gli3 protein processing to generate the Gli3 repressor is mediated by proteasome and inhibited by Hedgehog signaling. The Gli3 repressor concentration is graded along the anterior,posterior axis of the developing vertebrate limb due to posteriorly restricted Sonic hedgehog expression. In this study, we created a small deletion at the Gli3 locus (Gli3,68), which causes a half reduction in the Gli3 repressor levels and a slightly increased activity of full-length mutant protein in the limb. Mice homozygous for Gli3,68 develop one to two extra partial digits in the anterior of the limb, while mice carrying one copy of the Gli3,68 allele die soon after birth and display seven digits. These phenotypes are more severe than those found in mice lacking one wild-type Gli3 allele. The expression of dHand, Hoxd12, and Hoxd13 is anteriorly expanded in the limb, even though no up-regulation of Gli1 and Ptc RNA expression is detected. These findings suggest that a decrease in the Gli3 repressor level in combination with an increase in Gli3 full-length activity results in more severe digit patterning abnormalities than those caused by a loss of one wild-type Gli3 allele. Developmental Dynamics 236:769,776, 2007. © 2007 Wiley-Liss, Inc. [source] Crystallization for the Downstream Processing of ProteinsENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 3 2005S. Schmidt Abstract Protein crystallization offers great potential in downstream processing of pharmaceutical protein active ingredients. The advantages, which are well known and widely utilized in low-molecular weight crystallization, can also be expected to be found to some extent in protein crystallization. However, there is still a marked need for improvement in two main areas of protein processing, namely, in crystallization from impure solutions and scale-up. [source] Course and outcome of hepatitis CHEPATOLOGY, Issue 5B 200231 Center Dr., Jay H. Hoofnagle Bldg. 3, Room 9A2 The hepatitis C virus (HCV) is a small enveloped RNA virus belonging to the family flaviviridae and genus hepacivirus. The HCV RNA genome is 9,600 nucleotides in length and encodes a single polyprotein that is post-translationally cleaved into 10 polypeptides including t3 structural (C, E1, and E2) and multiple nonstructural proteins ([NS] NS2 to NS5). The NS proteins include enzymes necessary for protein processing (proteases) and viral replication (RNA polymerase). The virus replicates at a high rate in the liver and has marked sequence heterogeneity. There are 6 genotypes and more than 90 subtypes of HCV, the most common in the United States being 1a and 1b (approximately 75%), 2a and 2b (approximately 15%), and 3 (approximately 7%). Acute hepatitis C is marked by appearance of HCV RNA in serum within 1 to 2 weeks of exposure followed by serum alanine aminotransferase (ALT) elevations, and then symptoms and jaundice. Antibody to HCV (anti-HCV) tends to arise late. In acute resolving hepatitis, HCV RNA is cleared and serum ALT levels fall to normal. However, 55% to 85% of patients do not clear virus, but develop chronic hepatitis C. Chronic hepatitis C is often asymptomatic, but is usually associated with persistent or fluctuating elevations in ALT levels. The chronic sequelae of hepatitis C include progressive hepatic fibrosis, cirrhosis, and hepatocellular carcinoma. Extra-hepatic manifestations include sicca syndrome, cryoglobulinemia, glomerulonephritis, and porphyria cutanea tarda. Knowledge of the course and outcome of hepatitis C is important in developing approaches to management and therapy. [source] Redox-based endoplasmic reticulum dysfunction in neurological diseasesJOURNAL OF NEUROCHEMISTRY, Issue 1 2008Gábor Bánhegyi Abstract The redox homeostasis of the endoplasmic reticulum lumen is characteristically different from that of the other subcellular compartments. The concerted action of membrane transport processes and oxidoreductase enzymes maintain the oxidized state of the thiol-disulfide and the reducing state of the pyridine nucleotide redox systems, which are prerequisites for the normal functions of the organelle. The powerful thiol-oxidizing machinery allows oxidative protein folding but continuously challenges the local antioxidant defense. Alterations of the cellular redox environment either in oxidizing or reducing direction affect protein processing and may induce endoplasmic reticulum stress and unfolded protein response. The activated signaling pathways attempt to restore the balance between protein loading and processing and induce apoptosis if the attempt fails. Recent findings strongly support the involvement of this mechanism in brain ischemia, neuronal degenerative diseases and traumatic injury. The redox changes in the endoplasmic reticulum are integral parts of the pathomechanism of neurological diseases, either as causative agents, or as complications. [source] Caspase cleavage of exon 9 deleted presenilin-1 is an early event in apoptosis induced by calcium ionophore A 23187 in SH-SY5Y neuroblastoma cellsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2001Bogdan O. Popescu Abstract Presenilins (PSs) are mutated in a majority of familial Alzheimer disease (FAD) cases. Mutated PSs may cause FAD by a number of pro-apoptotic mechanisms, or by regulating ,-secretase activity, a protease involved in ,-amyloid precursor protein processing to the neurotoxic ,-amyloid peptide. Besides their normal endoproteolytic processing, PSs are substrates for caspases, being cleaved to alternative N-terminal and C-terminal fragments. So far little is known about the role of PSs cleavage in the apoptotic machinery. Here, we used SH-SY5Y neuroblastoma cells stably transfected with wild-type or exon 9 deleted presenilin 1 (PS1) in a time-course study after the exposure to the calcium ionophore A23187. During and after exposure to A 23187, intracellular calcium levels were higher in exon 9 deleted PS1 cells as compared with non-transfected and wild-type PS1 transfected cells. Cell death and the enrichment of apoptotic cells after A23187 exposure were increased by overexpression of exon 9 deleted PS1 as compared with the control cell lines. Wild-type PS1 cells were compared with exon 9 deleted PS1 cells and the temporal relationship between PS1 and other caspase substrates cleavages was analyzed. Exon 9 deleted PS1 cells exhibited a higher caspase-3 activation and a greater cleavage of PS1 and poly(ADP-ribose) polymerase (PARP) compared with wild-type PS1 cells. Exon 9 deleted PS1 cleavage occurred earlier than other caspase substrate cleavages (i.e., PARP and gelsolin), simultaneous with minimum detectable caspase-3 activation. Therefore, alternative cleavage of PS1 may play an important role for the regulation of the proteolytic cascade activated during apoptosis. J. Neurosci. Res. 66:122,134, 2001. © 2001 Wiley-Liss, Inc. [source] Unconventional serine proteases: Variations on the catalytic Ser/His/Asp triad configurationPROTEIN SCIENCE, Issue 12 2008an Ekici, Özlem Do Abstract Serine proteases comprise nearly one-third of all known proteases identified to date and play crucial roles in a wide variety of cellular as well as extracellular functions, including the process of blood clotting, protein digestion, cell signaling, inflammation, and protein processing. Their hallmark is that they contain the so-called "classical" catalytic Ser/His/Asp triad. Although the classical serine proteases are the most widespread in nature, there exist a variety of "nonclassical" serine proteases where variations to the catalytic triad are observed. Such variations include the triads Ser/His/Glu, Ser/His/His, and Ser/Glu/Asp, and include the dyads Ser/Lys and Ser/His. Other variations are seen with certain serine and threonine peptidases of the Ntn hydrolase superfamily that carry out catalysis with a single active site residue. This work discusses the structure and function of these novel serine proteases and threonine proteases and how their catalytic machinery differs from the prototypic serine protease class. [source] Global gene expression profile of Orientia tsutsugamushiPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 8 2010Bon-A Cho Abstract Orientia tsutsugamushi, an obligate intracellular bacterium, is the causative agent of Scrub typhus. The control mechanisms for bacterial gene expression are largely unknown. Here, the global gene expression of O. tsutsugamushi within eukaryotic cells was examined using a microarray and proteomic approaches for the first time. These approaches identified 643 genes, corresponding to approximately 30% of the genes encoded in the genome. The majority of expressed genes belonged to several functional categories including protein translation, protein processing/secretion, and replication/repair. We also searched the conserved sequence blocks (CSBs) in the O. tsutsugamushi genome which is unique in that up to 40% of its genome consists of dispersed repeated sequences. Although extensive shuffling of genomic sequences was observed between two different strains, 204 CSBs, covering 48% of the genome, were identified. When combining the data of CSBs and global gene expression, the CSBs correlates well with the location of expressed genes, suggesting the functional conservation between gene expression and genomic location. Finally, we compared the gene expression of the bacteria-infected fibroblasts and macrophages using microarray analysis. Some major changes were the downregulation of genes involved in translation, protein processing and secretion, which correlated with the reduction in bacterial translation rates and growth within macrophages. [source] Towards identifying Brassica proteins involved in mediating resistance to Leptosphaeria maculans: A proteomics-based approachPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 17 2008Nidhi Sharma Abstract To better understand the pathogen-stress response of Brassica species against the ubiquitous hemi-biotroph fungus Leptosphaeria maculans, we conducted a comparative proteomic analysis between blackleg-susceptible Brassica napus and blackleg-resistant Brassica carinata following pathogen inoculation. We examined temporal changes (6, 12, 24, 48 and 72,h) in protein profiles of both species subjected to pathogen-challenge using two-dimensional gel electrophoresis. A total of 64 proteins were found to be significantly affected by the pathogen in the two species, out of which 51 protein spots were identified using tandem mass spectrometry. The proteins identified included antioxidant enzymes, photosynthetic and metabolic enzymes, and those involved in protein processing and signaling. Specifically, we observed that in the tolerant B. carinata, enzymes involved in the detoxification of free radicals increased in response to the pathogen whereas no such increase was observed in the susceptible B. napus. The expression of genes encoding four selected proteins was validated using quantitative real-time PCR and an additional one by Western blotting. Our findings are discussed with respect to tolerance or susceptibility of these species to the pathogen. [source] Proteome analysis of antibody-producing CHO cell lines with different metabolic profilesBIOTECHNOLOGY & BIOENGINEERING, Issue 2 2007Deborah E. Pascoe Abstract Two-dimensional gel electrophoresis and tandem mass spectrometry were used to identify proteins associated with a metabolic shift during fed-batch cultures of two recombinant antibody-producing CHO cell lines. The first cell line underwent a marked change in lactate metabolism during culture, initially producing lactate and then consuming it, while the second cell line produced lactate for a similar duration but did not later consume it. The first cell line displayed a declining specific antibody productivity during culture, correlating to the 2-D gel results and the intracellular antibody concentration determined by HPLC. Several statistical analysis methods were compared during this work, including a fixed fold-change criterion and t -tests using standard deviations determined in several ways from the raw data and mathematically transformed data. Application of a variance-stabilizing transformation enabled the use of a global empirical standard deviation in the t -tests. Most of the protein spots changing in each cell line did not change significantly in the other cell line. A substantial fraction of the changing proteins were glycolytic enzymes; others included proteins related to antibody production, protein processing, and cell structure. Enolase, pyruvate kinase, BiP/GRP78, and protein disulfide isomerase were found in spots that changed over time in both cell lines, and some protein changes differed from previous reports. These data provide a foundation for future investigation of metabolism in industrially relevant mammalian cell culture processes, and suggest that along with differences between cell types, the proteins expressed in cultures with low lactate concentrations may depend on how those conditions were generated. Biotechnol. Bioeng. 2007;98: 391,410. © 2007 Wiley Periodicals, Inc. [source] A PCR-based strategy to generate yeast strains expressing endogenous levels of amino-terminal epitope-tagged proteinsBIOTECHNOLOGY JOURNAL, Issue 4 2008Keith R. Booher Abstract An epitope tag introduced to a gene of interest (GOI) greatly increases the ease of studying cellular proteins. Rapid PCR-based strategies for epitope tagging a protein's C-terminus at its native gene locus are widely used in yeast. C-terminal epitope tagging is not suitable for all proteins, however. Epitope tags fused to the C-terminus can interfere with function of some proteins or can even be removed by C-terminal protein processing. To overcome such problems, proteins can be tagged with epitopes at their amino-termini, but generating yeast strains expressing N-terminal epitope tagged genes under control of the endogenous promoter at the native locus is comparatively more difficult. Strategies to introduce N-terminal epitope tags have been reported previously but often introduce additional sequences other than the epitope tag into the genome. Furthermore, N-terminal tagging of essential genes by current methods requires formation of diploid strains followed by tetrad dissection or expression of an additional copy of the GOI from a plasmid. The strategies described here provide a quick, facile means of epitope tagging the N-terminus of both essential and nonessential genes in a two-step PCR-based procedure. The procedure has the significant advantage of leaving tagged genes under the control of their endogenous promoters, and no additional sequences other than the epitope tag encoding nucleotides are inserted into the genome. [source] | ||||||||||||||||||||||||||||