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- Agaton, C., et al.
(författare)
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Affinity proteomics for systematic protein profiling of chromosome 21 gene products in human tissues
- 2003
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 2, s. 405-
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Tidskriftsartikel (refereegranskat)abstract
- Here we show that an affinity proteomics strategy using affinity-purified antibodies raised against recombinant human protein fragments can be used for chromosome-wide protein profiling. The approach is based on affinity reagents raised toward bioinformatics-designed protein epitope signature tags corresponding to unique regions of individual gene loci. The genes of human chromosome 21 identified by the genome efforts were investigated, and the success rates for de novo cloning, protein production, and antibody generation were 85, 76, and 56%, respectively. Using human tissue arrays, a systematic profiling of protein expression and subcellular localization was undertaken for the putative gene products. The results suggest that this affinity proteomics strategy can be used to produce a proteome atlas, describing distribution and expression of proteins in normal tissues as well as in common cancers and other forms of diseased tissues.
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| 4. |
- Huang, Fang, et al.
(författare)
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Isolation of Outer Membrane of Synechocystis sp. PCC 6803 and Its Proteomic Characterization
- 2004
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Ingår i: Molecular & Cellular Proteomics. - : American Society for Biochemistry and Molecular Biology. - 1535-9476 .- 1535-9484. ; 3:6, s. 586-595
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Tidskriftsartikel (refereegranskat)abstract
- In this report, we describe a newly developed method for isolating outer membranes from Synechocystis sp. PCC 6803 cells. The purity of the outer membrane fraction was verified by immunoblot analysis using antibodies against membrane-specific marker proteins. We investigated the protein composition of the outer membrane using two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry followed by database identification. Forty-nine proteins were identified corresponding to 29 different gene products. All of the identified proteins have a putative N-terminal signal peptide. About 40% of the proteins identified represent hypothetical proteins with unknown function. Among the proteins identified are a Toc75 homologue, a protein that was initially found in the outer envelope of chloroplasts in pea, as well as TolC, putative porins, and a pilus protein. Other proteins identified include ABC transporters and GumB, which has a suggested function in carbohydrate export. A number of proteases such as HtrA were also found in the outer membrane of Synechocystis sp. PCC 6803.
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| 5. |
- Huang, F, et al.
(författare)
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Proteomics of Synechocystis sp strain PCC 6803 - Identification of plasma membrane proteins
- 2002
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 1:12, s. 956-966
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Tidskriftsartikel (refereegranskat)abstract
- Cyanobacteria are unique prokaryotes since they in addition to outer and plasma membranes contain the photosynthetic membranes (thylakoids) The plasma membranes of Synechocystis 6803, which can be completely purified by density centrifugation and polymer two-phase partitioning, have been found to be more complex than previously anticipated, i. e they appear to be essential for assembly of the two photosystems. A proteomic approach for the characterization of cyanobacterial plasma membranes using two-dimensional gel electrophoresis and mass spectrometry analysis revealed a total of 57 different membrane proteins of which 17 are integral membrane spanning proteins. Among the 40 peripheral proteins 20 are located on the periplasmic side of the membrane, while 20 are on the cytoplasmic side. Among the proteins identified are subunits of the two photosystems as well as Vipp1, which has been suggested to be involved in vesicular transport between plasma and thylakoid membranes and is thus relevant to the possibility that plasma membranes are the initial site for photosystem biogenesis. Four subunits of the Pilus complex responsible for cell motility were also identified as well as several subunits of the TolC and TonB transport systems. Several periplasmic and ATP-binding proteins of ATP-binding cassette transporters were also identified as were two subunits of the F-0 membrane part of the ATP synthase
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| 10. |
- Malmström, Johan, et al.
(författare)
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Transforming growth factor-beta 1 specifically induce proteins involved in the myofibroblast contractile apparatus
- 2004
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Ingår i: Molecular & Cellular Proteomics. - 1535-9484. ; 3:5, s. 466-477
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Tidskriftsartikel (refereegranskat)abstract
- Transforming growth factor-beta(1) (TGF-beta(1)) induces alpha-smooth muscle actin (alpha-SMA) and collagen synthesis in fibroblast both in vivo and in vitro and plays a significant role in tissue repair and the development of fibrosis. During these processes the fibroblasts differentiate into activated fibroblasts (so called myofibroblasts), characterized by increased alpha-SMA expression. Because TGF-beta(1) is considered the main inducer of the myofibroblast phenotype and cytoskeletal changes accompany this differentiation, the main objective of this investigation was to study how TGF-beta(1) alters protein expression of cytoskeletal-associated proteins. Metabolic labeling of cell cultures by [(35)S]methionine, followed by protein separation on two-dimensional gel electrophoresis, displayed approximately 2500 proteins in the pI interval of 3-10. Treatment of TGF-beta(1) led to specific spot pattern changes that were identified by mass spectrometry and represent specific induction of several members of the contractile apparatus such as calgizzarin, cofilin, and profilin. These proteins have not previously been shown to be regulated by TGF-beta(1), and the functional role of these proteins is to participate in the depolymerization and stabilization of the microfilaments. These results show that TGF-beta(1) induces not only alpha-SMA but a whole set of actin-associated proteins that may contribute to the increased contractile properties of the myofibroblast. These proteins accompany the induced expression of alpha-SMA and may participate in the formation of stress fibers, cell contractility, and cell spreading characterizing the myofibroblasts phenotype.
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